Melatonin decreases in vitro viability and migration of spheres derived from CF41.Mg canine mammary carcinoma cells.

BACKGROUND: Mammary cancer is a common disease affecting female dogs, where approximately 50% of the cases are malignant. There is a subpopulation of cancer cells with stem cell-like features within the tumour microenvironment, which can form in vitro spheres, cell structures that grow in anchor-free conditions. This cell population shows resistance to conventional antitumor treatments explaining in part the recurrence of some type of cancers. It has been previously reported that spheres derived from CF41.Mg canine mammary carcinoma cells exhibit several stemness features. Melatonin has shown antitumor effects on cancer mammary cells; nevertheless, its effects have been poorly evaluated on canine mammary cancer stem-like cells. In this regard, it has described that melatonin decreases the expression of OCT-4 in CMT-U2229 mammary cancer cells, a transcription factor that participates in the modulation of self-renewal and drug resistance in cancer stem-like cells. The aim of this study was to compare the effects of melatonin on viability and migration of canine mammary carcinoma CF41.Mg-spheres, and CF41.Mg-parental cells. CF41.Mg cells were grown in DMEM high-glucose medium containing 10% bovine foetal serum. CF41.Mg-spheres were cultured in ultra-low attachment plates with serum-free DMEM/F12 containing several growth factors. Cell viability (MTS reduction) and migration (transwell) assays were conducted in presence of melatonin (0.01, 0.1 or 1 mM). RESULTS: Melatonin decreased cell viability at 1 mM (P < 0.05), with a significant reduction in spheres compared to parental cells at 24 and 48 h (P < 0.05). Cell migration was inhibited in response to non-cytotoxic concentration of melatonin (0.1 mM) (P < 0.05) in spheres and monolayer of cells, no significant differences were detected between both cell subtypes. CONCLUSIONS: These results indicate that melatonin reduces viability and migration of CF41.Mg cells, where spheres exhibit greater sensitivity to the hormone. Thus, melatonin represents a valuable potential agent against mammary cancer cells, especially cancer stem-like cells.

Carcinogenesis: the cancer cell-mast cell connection.

BACKGROUND: In mammals, inflammation is required for wound repair and tumorigenesis. However, the events that lead to inflammation, particularly in non-healing wounds and cancer, are only partly understood. FINDINGS: Mast cells, due to their great plasticity, could orchestrate the inflammatory responses inducing the expression of extraembryonic programs of normal and pathological tissue formation. This heterogeneity of mast cells could allow a microenvironment to be recreated similar to the extraembryonic structures, i.e., amnion and yolk sac, which are needed for embryonic development. Mast cells could provide a framework for understanding the connection between inflammation and tumor growth, invasion and metastasis. In this way, the mast cells could express inflammatory phenotypes, which would enable the cancer stem cells to develop. Thus, the cancer cell uses mast cells to express the extraembryonic functions that are needed to allow the cancer stem cell to proliferate and invade. If so, then by using this appropriate inflammatory interstitial microenvironment, a cancer stem cell can reach maximum levels of growth and invasion inside the host. CONCLUSION: Therefore, the comparison of tumors with wounds that do not heal would be supported since both pathological processes use extraembryonic mechanisms by mast cells. The adoption of these mechanisms warrants tumor survival in an embryonic-like state.

Different Trypanosoma cruzi calreticulin domains mediate migration and proliferation of fibroblasts in vitro and skin wound healing in vivo.

Calreticulin is an endoplasmic reticulum-resident, calcium-binding, stress-produced, chaperone protein that serves multiple functions and is widely distributed in eukaryotic cells. Exogenously applied recombinant calreticulin solution, markedly enhanced the rate and quality of skin wound healing. These modulatory effects are more efficient than commercially available topic platelet-derived growth factor ointments (Regranex®). Trypanosoma cruzi calreticulin is more effective in equimolar terms to human counterpart in accelerating skin wound healing. While the effect of externally added recombinant parasite calreticulin on wound healing has been reported, the domains responsible for these modulatory effects have not yet been established. Here, recombinant parasite calreticulin and some of its domains were tested to assess their influence in increasing proliferation and migration of fibroblasts in vitro and rat skin wound healing in vivo. Herein, we propose that Trypanosoma cruzi whole calreticulin or some of its domains are differentially involved in the modulation of wound-healing cell migration and proliferation, and cosmetic outcome. Therefore, precise combination of the parasite protein and its domains could allow us to tailor-specific desired effects during the skin wound-healing process.

MicroRNA deregulation in triple negative breast cancer reveals a role of miR-498 in regulating BRCA1 expression.

Emerging evidence suggests that BRCA1 pathway contributes to the behavior of sporadic triple negative breast cancer (TNBC), but little is known about the mechanisms underlying this association. Considering the central role that microRNAs (miRNAs) play in gene expression regulation, the aim of this study was to identify miRNAs specifically deregulated in TNBC and investigate their involvement in BRCA1 regulation. Using locked nucleic acid (LNA)-based microarrays, expression levels of 1919 miRNAs were measured in paraffin-embedded tissues from 122 breast tumors and 11 healthy breast tissue samples. Differential miRNA expression was explored among the main subtypes of breast cancer, and 105 miRNAs were identified as specific for triple negative tumors. In silico prediction revealed that miR-498 and miR-187-5p target BRCA1, and these results were confirmed by luciferase reporter assay. While miR-187-5p was found overexpressed in a luminal B cell line, miR-498 was highly expressed in a triple negative cell line, Hs578T, and its expression was negatively correlated with the levels of BRCA1. We functionally demonstrated that miR-498 inhibits BRCA1 in breast cancer cell lines, and showed that inhibition of miR-498 led to reduced proliferation in the triple negative cell line Hs578T. Our results indicate that miR-498 regulates BRCA1 expression in breast cancer and its overexpression could contribute to the pathogenesis of sporadic TNBC via BRCA1 downregulation.

Breastfeeding and Immunohistochemical Expression of ki-67, p53 and BCL2 in Infiltrating Lobular Breast Carcinoma.

BACKGROUND/AIM: Invasive lobular breast carcinoma is the second most common type of breast cancer after invasive ductal carcinoma. According to the American Cancer Society, more than 180,000 women in the United States find out they have invasive breast cancer each year. Personal history of breast cancer and certain changes in the breast are correlated with an increased breast cancer risk. The aim of this work was to analyze breastfeeding in patients with infiltrating lobular breast carcinoma, in relation with: 1) clinicopathological parameters, 2) hormonal receptors and 3) tissue-based tumor markers. MATERIALS AND METHODS: The study included 80 women with ILC, 46 of which had breastfeed their children. Analyzed parameters were: age, tumor size, axillary lymph node (N), distant metastasis (M), histological grade (HG), estrogen receptor (ER), progesterone receptor (PR), androgen receptor (AR), Ki-67, p53 and BCL2. RESULTS: ILC of non-lactating women showed a larger (p = 0.009), lymph node involvement (p = 0.051) and distant metastasis (p = 0.060). They were also more proliferative tumors measured by Ki-67 (p = 0.053). Breastfeeding history did not influence the subsequent behavior of the tumor regardless of histological subtype. CONCLUSION: Lactation seems to influence the biological characteristics of ILC defining a subgroup with more tumor size, axillary lymph node involvement, distant metastasis and higher proliferation measured by ki-67 expression.

Tumor microRNA expression profiling identifies circulating microRNAs for early breast cancer detection.

BACKGROUND: The identification of novel biomarkers for early breast cancer detection would be a great advance. Because of their role in tumorigenesis and stability in body fluids, microRNAs (miRNAs) are emerging as a promising diagnostic tool. Our aim was to identify miRNAs deregulated in breast tumors and evaluate the potential of circulating miRNAs in breast cancer detection. METHODS: We conducted miRNA expression profiling of 1919 human miRNAs in paraffin-embedded tissue from 122 breast tumors and 11 healthy breast tissue samples. Differential expression analysis was performed, and a microarray classifier was generated. The most relevant miRNAs were analyzed in plasma from 26 healthy individuals and 83 patients with breast cancer (36 before and 47 after treatment) and validated in 116 healthy individuals and 114 patients before treatment. RESULTS: We identified a large number of miRNAs deregulated in breast cancer and generated a 25-miRNA microarray classifier that discriminated breast tumors with high diagnostic sensitivity and specificity. Ten miRNAs were selected for further investigation, of which 4 (miR-505-5p, miR-125b-5p, miR-21-5p, and miR-96-5p) were significantly overexpressed in pretreated patients with breast cancer compared with healthy individuals in 2 different series of plasma. MiR-505-5p and miR-96-5p were the most valuable biomarkers (area under the curve 0.72). Moreover, the expression levels of miR-3656, miR-505-5p, and miR-21-5p were decreased in a group of treated patients. CONCLUSIONS: Circulating miRNAs reflect the presence of breast tumors. The identification of deregulated miRNAs in plasma of patients with breast cancer supports the use of circulating miRNAs as a method for early breast cancer detection.

Histological grade (HG) in invasive ductal carcinomas of the breast of less than 1 cm: clinical and biological associations during progression from HG1 to HG3.

AIM: To study the clinical and biological (cellular proliferation and hormone-dependence) associations during the progression of histological grade (HG), from HG1 to HG3, in invasive ductal carcinomas of the breast (IDC) <1 cm. PATIENTS AND METHODS: The study group included 119 women with IDCs ≤1 cm, aged between 27 and 88 years (median=61 years). The parameters analyzed were: histological grade (HG1: 52; HG2: 45; HG3: 22); axillary lymph node involvement (N); distant metastasis (M); and immunohistochemical expression of estrogen (ER), progesterone (PR) and androgen (AR) receptors, and Ki67, p53 and B-cell lymphoma 2 (BCL2). RESULTS: Compared to HG3 tumors, HG1s exhibited an increased expression of ER, AR and BCL2, as well as lower expression of p53 and Ki67. In HG1 tumors, significant (p<0.05) associations were found between ER and PR (positive), ER and p53 (negative), ER and Ki67 (negative), PR and AR (positive), PR and p53 (negative), AR and p53 (negative), p53 and BCL2 (negative), and between BCL2 and Ki67 (negative). HG3s only showed significant (p<0.05) associations between ER and Ki-67 (negative) and between BCL2 or Ki-67 (negative). Only two significant relationships (ER-Ki67 and BCL2-Ki67) persisted in all three grades. CONCLUSION: Our results lead us to the following conclusions: i) compared HG1, HG3 ductal carcinomas exhibited decreased expression of ER, AR and BCL2 and increased expression of p53 and Ki67; and ii) only two significant and negative relations (ER-Ki67 and BCL2-Ki67) persisted in all three grades.

Clinicopathological characteristics of infiltrating lobular breast carcinoma in elderly women: Preliminary results.

This study was conducted to investigate the clinicopathological parameters in elderly women (aged >70 years) with infiltrating lobular carcinoma (ILC) of the breast and compare the results with those obtained from younger patients (aged 55-70 years). The study sample included a total of 46 women with ILCs, 10 aged >70 and 36 aged 55-70 years. The parameters analysed were tumor size, histological grade (HG), axillary lymph node involvement, distant metastasis and immunohistochemical expression of estrogen, progesterone and androgen receptors, Ki67, p53 and B cell lymphoma 2. Compared to women aged 55-70 years, ILCs in women aged >70 years were commonly of larger size (P=0.068) and were more frequently HG3 (P=0.024). There were no statistically significant differences in the other parameters analysed. Furthermore, we were unable to determine differences in cancer recurrence and mortality in the two patient subgroups during our follow-up. In conclusion, our preliminary results, based on the limited number of cases included in this study, indicate that i) ILCs in women aged >70 years tended to be larger compared to those in women aged 55-70 years and were more frequently of grade 3; and ii) there were no significant differences in terms of recurrence and mortality between the two patient subgroups during our follow-up.

CA15.3 serum concentrations in older women with infiltrating ductal carcinomas of the breast.

Breast cancer is currently becoming a disease of the elderly. We have studied the relation between CA 15.3 serum concentrations and clinical-pathological parameters in 69 women with IDC aged over 70 years (76.3±4.2; range: 71-88; median 76). A group of 205 women with the same tumor but aged <70 years (62.8±4.0; range: 55-70; median 63) was also considered for comparison. Tumor size, axillary lymph node involvement, distant metastasis and histological grade were taken account. Serum CA 15.3 was determined by luminescence assay. CA 15.3 serum concentrations ranged between 6 and 85 U/mL (median 22.9 U/mL), and were higher only in patients with greater (qualitative and quantitative; p: 0.041) tumor size. Our results show that in women with IDCs, and aged over 70 years, serum CA 15.3 serum concentrations are associated exclusively with a greater tumor size, being these findings different to those described in women with the same subtype of tumor considered as a whole or with lower age.

Wound healing reaction: A switch from gestation to senescence.

The repair of wounded tissue during postnatal life could be associated with the upregulation of some functions characteristic of the initial phases of embryonic development. The focusing of these recapitulated systemic functions in the interstitial space of the injured tissue is established through a heterogeneous endothelial barrier which has excretory-secretory abilities which in turn, would induce a gastrulation-like process. The repair of adult tissues using upregulated embryonic mechanisms could explain the universality of the inflammatory response against injury, regardless of its etiology. However, the early activation after the injury of embryonic mechanisms does not always guarantee tissue regeneration since their long-term execution is mediated by the host organism.

Positive immunohistochemical expression of bcl-2 in hormone-independent breast carcinomas is associated with a greater lymph node involvement and poor outcome.

To study the immunohistochemical expression of bcl-2 in patients with hormone-independent breast infiltrating ductal carcinomas (IDC) and its possible association with other clinico-biological parameters and outcome. Our study group included 72 females with hormone-independent (ER and PgR negative) infiltrating ductal breast carcinomas. Age, tumor size, axillary lymph node involvement (N), distant metastasis and histological grade, as well as the immunohistochemical expression of Ki67, p53 and androgen receptor (AR), were analyzed. We follow up 57 patients during a period of time ranged between 20 and 193 months (80.2 ± 58.3; median 78 months). Of all IDCs included in our study, 18 were ER-/PgR-/bcl-2+ and 54 ER-/PgR-/bcl-2-. The percentages of slightly bcl-2-positive (+) and bcl-2-strong positive (++) cases were 25 and 19 %, respectively, values lower than those observed in ER+/PgR+ tumors (79.3 and 86.8 %, respectively). Breast IDC with positivity (+) for bcl-2 showed, exclusively, greater lymph node involvement higher than 3 nodes (N+ >3) (p 0.021) and a great number of deaths due to the tumor (p 0.011). Same results were obtained when we compared bcl-2-negative and bcl-2-strong positive (++) subgroups. Our results led us to consider that the positive (+ or ++) immunohistochemical expression of bcl-2 in hormone-independent (ER and PgR negative) breast carcinomas is associated with greater axillary lymph node involvement and a greater number of deaths in the follow-up, being these data opposite to that observed in hormone-dependent tumors.

Association between tumor size and immunohistochemical expression of Ki-67, p53 and BCL2 in a node-negative breast cancer population selected from a breast cancer screening program.

BACKGROUND/AIM: Breast cancer is the most common type of cancer among women. Breast infiltrating ductal carcinoma (IDC) is the most common type of breast cancer, approximately 80% of all breast carcinomas. The aim of this study was to analyze the association of tumor size, evaluated after histopathological analysis, with different clinical and biological parameters in IDC. MATERIALS AND METHODS: The study group included 251 women with IDC without axillary lymph node involvement, aged between 27 and 81 years. Analyzed parameters were: age, histological grade, menopausal status, menarche, pregnancy, abortion, breastfeeding, contraceptive use, hormone replacement therapy, estrogen receptor (ER), progesterone receptor (PR), androgen receptor (AR), Ki-67, p53 and BCL2. RESULTS: Pathological tumor size was between 0.2 and 5.1 cm (1.43±0.86 cm). Tumors in 45 cases exceeded 2 cm, in eight 3 cm and only in one 5 cm. Pathological size was significantly associated with age >70 vs. <50 years (p=0.054), histological grade III vs. I (p=0.0003), positivity for Ki-67 (p=0.0003) and for p53 (p=0.0032). CONCLUSION: Tumor size was significantly associated with age >70 years, histological grade 3 and immunohistochemically-augmented expression of Ki-67 and p53.

Surgical inflammatory stress: the embryo takes hold of the reins again.

The surgical inflammatory response can be a type of high-grade acute stress response associated with an increasingly complex trophic functional system for using oxygen. This systemic neuro-immune-endocrine response seems to induce the re-expression of 2 extraembryonic-like functional axes, i.e. coelomic-amniotic and trophoblastic-yolk-sac-related, within injured tissues and organs, thus favoring their re-development. Accordingly, through the up-regulation of two systemic inflammatory phenotypes, i.e. neurogenic and immune-related, a gestational-like response using embryonic functions would be induced in the patient's injured tissues and organs, which would therefore result in their repair. Here we establish a comparison between the pathophysiological mechanisms that are produced during the inflammatory response and the physiological mechanisms that are expressed during early embryonic development. In this way, surgical inflammation could be a high-grade stress response whose pathophysiological mechanisms would be based on the recapitulation of ontogenic and phylogenetic-related functions. Thus, the ultimate objective of surgical inflammation, as a gestational process, is creating new tissues/organs for repairing the injured ones. Since surgical inflammation and early embryonic development share common production mechanisms, the factors that hamper the wound healing reaction in surgical patients could be similar to those that impair the gestational process.

The wound-healing response and upregulated embryonic mechanisms: brothers-in-arms forever.

The cutaneous wound-healing reaction occurs in overlapping but inter-related phases, which ultimately result in fibrosis. The pathophysiological mechanisms involved in fibrotic diseases, including organ-related and even systemic diseases, such as systemic sclerosis, could represent the successive systemic upregulation of extraembryonic-like phenotypes, that is, amniotic and vitelline phenotypes. These two extraembryonic-like phenotypes act on the injured tissue to induce a process similar to gastrulation, which occurs during the early phases of embryo development. The amniotic-like phenotype plays a leading role in the development of neurogenic responses with significant hydroelectrolytic alterations that essentially represent the development of open microcirculation within the injured tissue. In turn, through the overlapping expression of a vitelline-like phenotype, a bone marrow-related response is produced. Interstitial infiltration by molecular and cellular mediators contributed by amniotic- and vitelline-like functions provides the functional and metabolic autonomy needed for inducing new tissue formation through mechanisms similar to those that act in gastrulation during the early phases of embryonic development. Thus, while a new tissue is formed, it quickly evolves into fibrotic tissue because of premature senescence. Mechanisms related to extraembryonic-like functions have been suggested in the following physiological and pathological processes: embryonic development; wound-healing reactions occurring during adult life; and senescence. The existence of this sort of basic self-organizing fractal-like functional pattern is an essential characteristic of our way of life.

Portal hypertensive cardiovascular pathology: the rescue of ancestral survival mechanisms?

The portal system derives from the vitelline system, which is an extra-embryonic venous system. It could be suggested that this extraembryonic origin determines some of the characteristics attributed to portal hypertension, both compensated, i.e. prehepatic, and decompensated, i.e. fibrotic or cirrhotic. The experimental models most frequently used for studying both types of portal hypertension are portal vein ligation and common bile duct ligation in rats, respectively. We propose that in partial portal vein ligated rats, a low-grade inflammatory response, formed by the successive expression of three overlapping phenotypes - ischemia-reperfusion, vitellogenic-like and remodeling or gastrulation-like - is produced. The names of these inflammatory phenotypes developed in compensated portal hypertension are based on some metabolic similarities that can be established with the abovementioned phases of embryonic development. In bile-duct ligated rats, decompensation related to hepatic insufficiency would induce a high-grade inflammatory response. In this experimental model, the splanchnic interstitium, the mesenteric lymph and the peritoneal mesothelium seem to create an inflammatory axis that produces ascites. The functional comparison between the ascitic and the amniotic fluids would imply that, in the decompensated portal hypertensive syndrome, the abdominal mesothelium acquires properties of the amniotic membranes or amnion. In conclusion, the hypothetical comparison between the inflammatory portal hypertensive evolutive types and the evolutive phases of embryonic development could allow for translational research.

The Amazing Power of Cancer Cells to Recapitulate Extraembryonic Functions: The Cuckoo's Tricks.

Inflammation is implicated in tumor development, invasion, and metastasis. Hence, it has been suggested that common cellular and molecular mechanisms are activated in wound repair and in cancer development. In addition, it has been previously proposed that the inflammatory response, which is associated with the wound healing process, could recapitulate ontogeny through the reexpression of the extraembryonic, that is, amniotic and vitelline, functions in the interstitial space of the injured tissue. If so, the use of inflammation by the cancer-initiating cell can also be supported in the ability to reacquire extraembryonic functional axes for tumor development, invasion, and metastasis. Thus, the diverse components of the tumor microenvironment could represent the overlapping reexpression of amniotic and vitelline functions. These functions would favor a gastrulation-like process, that is, the creation of a reactive stroma in which fibrogenesis and angiogenesis stand out.

CA 15-3 serum levels in patients with ductal breast carcinoma: relationship with clinicopathological parameters and tumor markers.

INTRODUCTION: Cancer antigen 15-3 (CA 15-3) is the most widely used serum marker in diagnosing and monitoring breast cancer. The aim of this work was to analyze preoperative CA 15-3 serum levels in patients with ductal breast carcinoma in relation to 1) clinicopathological parameters, 2) hormone receptors, and 3) tissue-based tumor markers. METHODS: A group of 340 women with infiltrating ductal carcinoma of the breast who had undergone no prior treatment was studied. These women ranged in age between 27 and 83 years (mean age 61.5±9.9 years). Preoperative CA 15-3 serum levels were determined by an immunoradiometric method. Hormone receptors (estrogen, progesterone and androgen receptors), p53, bcl-2 and Ki67 were determined by different immunohistochemical methods. Epidermal growth factor receptor (EGFR) was analyzed in the cell membranes by radioligand assay whereas cathepsin D and pS2 were determined by immunoradiometric analysis. Tumor ploidy and S-phase fraction were studied by flow cytometry. RESULTS: CA 15-3 serum levels were higher in postmenopausal women (p=0.032), in patients with tumors exceeding 2 cm (p=0.003), lymph node involvement (p=0.026), distant metastases (M1) (p<0.0001), S-phase fraction <7% (p=0.015), EGFR <6 fmol/mg protein (p=0.025), and cathepsin D <50 pmol/mg protein (p=0.023). CONCLUSIONS: Preoperative CA 15-3 serum levels were associated with cell proliferation determined by the S-phase fraction, the concentration of cathepsin D in the cytosol, and the EGFR concentration in the cell membrane.

Clinical and Biological Significance of Cathepsin D Levels in Breast Cancer Cytosol in Women Over 70 years.

OBJECTIVE: To study cytosolic cathepsin D behavior and possible relationship with other clinical and biological parameters in women affected by breast invasive ductal carcinomas and older than 70 years (range: 71-88). MATERIAL AND METHODS: cytosolic levels of cathepsin D were determined by an Immunoradiometric Assay (IRMA-CIS France). Clinical and biological factors analyzed were: size, axillary lymph node involvement, distant metastasis, histological grade, ploidy, S phase cell, cytosolic estrogen receptor, progesterone receptor and pS2, and concentrations of epidermal growth factor receptor (EGFR) in cell membranes. RESULTS: Cathepsin D concentrations ranged between 13 and 1228 pmol/mg prot.. Median value of 41 was considered as threshold of positivity. Cathepsin D positive tumors showed higher S-phase values (P = 0.046) and were most often histological grade III (P = 0.047). However, the most important finding was the existence of a positive correlation (r = 0.51786) and statistically significant (P < 0.05) between S-phase values and cathepsin D in the overall group of tumors, and those ER+, but not in ER-. We determined cathepsin D concentrations in 131 women with invasive ductal breast carcinomas, but aged between 50 and 70 years (median 61) and we did not find differences based on those values in women >70 years. In addition, we found no correlation between S-phase values and Cathepsin D, both overall and in relation with hormone dependence (ER). CONCLUSIONS: THOSE RESULTS LED US TO THE FOLLOWING CONCLUSIONS: (1) cytosolic concentrations of cathepsin D in invasive infiltrating breast carcinomas in women over 70 are similar to those seen in women with the same type of tumor, but aged 50 to 70 years and are associated with increased cell proliferation measured by S phase, and histological grade III; (2) in women older than 70 years, cathepsin D concentrations are statistically significantly correlated with phase synthesis values in hormone-dependent tumors, but not in hormone-independent, fact not observed in infiltrating ductal breast carcinomas of women aged between 50 and 70. This could reflect a different mitogenic role of the aspartyl protease enzyme linked to hormone dependence as age function parameter.

7q21-rs6964587 and breast cancer risk: an extended case-control study by the Breast Cancer Association Consortium.

BACKGROUND: Using the Breast Cancer Association Consortium, the authors previously reported that the single nucleotide polymorphism 7q21-rs6964587 (AKAP9-M463I) is associated with breast cancer risk. The authors have now assessed this association more comprehensively using 16 independent case-control studies. METHODS: The authors genotyped 14,843 invasive case patients and 19,852 control subjects with white European ancestry and 2595 invasive case patients and 2192 control subjects with Asian ancestry. ORs were estimated by logistic regression, adjusted for study. Heterogeneity in ORs was assessed by fitting interaction terms or by subclassifying case patients and applying polytomous logistic regression. RESULTS: For white European women, the minor T allele of 7q21-rs6964587 was associated with breast cancer risk under a recessive model (OR 1.07, 95% CI 1.00 to 1.13, p = 0.04). Results were inconclusive for Asian women. From a combined analysis of 24 154 case patients and 33,376 control subjects of white European ancestry from the present and previous series, the best-fitting model was recessive, with an estimated OR of 1.08 (95% CI 1.03 to 1.13, p = 0.001). The OR was greater at younger ages (p trend = 0.01). CONCLUSION: This may be the first common susceptibility allele for breast cancer to be identified with a recessive mode of inheritance.