We have defined a human breast tumor associated antigen using a murine monoclonal antibody (MAb DF3) prepared against a membrane-enriched fraction of a human breast carcinoma. This antigen has a MW of 290 kD and is detectable on the cell surface of human breast carcinoma cells using a live cell radioimmunoassay and fluorescence flow cytometry. More important, immunoperoxidase staining with MAb DF3 clearly distinguishes malignant and benign breast lesions. A cytoplasmic staining pattern has been observed with 40 of 51 (78%) breast carcinomas, but only one of 13 fibroadenoma or fibrocystic disease specimens. In contrast, reactivity of benign breast lesions with MAb DF3 primarily occurs along apical borders on ductules. These results demonstrate that the DF3 antigen is present on apical borders of more differentiated secretory mammary epithelial cells and in the cytosol of less differentiated cells.

The murine monoclonal antibody (mAb) DF3 reacts with a high molecular weight glycoprotein detectable in human breast carcinomas. DF3 antigen expression correlates with human breast tumor differentiation, and the detection of a cross-reactive species in human milk has suggested that this antigen might be useful as a marker of differentiated mammary epithelium. To further characterize DF3 antigen expression, we have isolated a cDNA clone from a Agtll library by screening with mAb DF3. The results demonstrate that this 309-base-pair cDNA, designated pDF9.3, codes for the DF3 epitope. Southern blot analyses of EcoRI-digested DNAs from six human tumor cell lines with 32P-labeled pDF9.3 have revealed a restriction fragment length polymorphism. Variations in size of the alleles detected by pDF9.3 were also identified in Pst I, but not in HindIII, DNA digests. Furthermore, hybridization of 32P-labeled pDF9.3 with total cellular RNA from each of these cell lines demonstrated either one or two transcripts that varied from 4.1 to 7.1 kilobases in size. The presence of differently sized transcripts detected by pDF9.3 was also found to correspond with the polymorphic expression of DF3 glycoproteins. Nucleotide sequence analysis of pDF9.3 has revealed a highly conserved (G+C)-rich 60-base-pair tandem repeat. These rmdings suggest that the variation in size of alleles coding for the polymorphic DF3 glycoprotein may represent different numbers of repeats.A human breast carcinoma-associated antigen has been identified by using a murine monoclonal antibody (mAb), designated DF3. mAb DF3 was prepared against a membrane-enriched fraction of a human breast carcinoma metastatic to liver (1). DF3 antigen is expressed on the apical borders of secretory mammary epithelial cells and in the cytosol of less differentiated malignant cells (1). DF3 antigen expression also correlates with the degree of breast tumor differentiation and estrogen receptor status (2). These findings and the detection of DF3 antigen in human milk (3) The present studies were performed to further examine genetic mechanisms responsible for the heterogeneity of DF3 antigen expression. We describe the isolation of a partial cDNA clone,t designated pDF9.3, coding for the DF3 antigen. This cDNA clone encompasses 309 nucleotides of DF3 mRNA and has tandemly repeated sequences. By using this clone, we also demonstrate that the heterogeneity of DF3 antigen is related to size variations in DF3 alleles and DF3 transcripts. MATERIALS AND METHODSLibrary Screening. An oligo(dT)-primed cDNA library prepared from human MCF-7 breast carcinoma cells in Agtll was kindly provided by P. Chambon (Institut de Chimie Biologique, Strasbourg, France) (8). Immunologic screening of the Agtll library was performed as described (9) by using affinity-purified mAb DF3 (0.25 ,ug/ml) and antimouse IgG conjugated with alkaline phosphatase (Promega Biotec, Madison, WI). Positive plaques were isolated and the phage was further purified to homogeneity by repeated antibody screening. DNA was i...

. Effects of menstrual cycle and physical training on heat loss responses during dynamic exercise at moderate intensity in a temperate environment. Am J Physiol Regul Integr Comp Physiol 288: R1347-R1353, 2005. First published January 27, 2005 doi:10.1152/ajpregu.00547.2004.-We evaluated the effects of the menstrual cycle and physical training on heat loss (sweating and cutaneous vasodilation) responses during moderate exercise in a temperate environment. Ten untrained (group U) and seven endurance-trained (group T) women (maximal O2 uptake of 36.7 Ϯ 1.1 vs. 49.4 Ϯ 1.7 ml ⅐ kg Ϫ1 ⅐ min Ϫ1 , respectively; P Ͻ 0.05) performed a cycling exercise at 50% maximal O 2 uptake for 30 min during both the midfollicular and midluteal menstrual phase in a temperate environment (ambient temperature of 25°C, relative humidity of 45%). In group U, plasma levels of estrone, estradiol, and progesterone at rest and esophageal temperature (Tes) during exercise were significantly higher during the midluteal than during the midfollicular phase (P Ͻ 0.05). Sweating rate and cutaneous blood flow (measured via laserDoppler flowmetry) on the chest, back, forearm, and thigh were lower during the midluteal than during the midfollicular phase during exercise. Tes threshold for heat loss responses was significantly higher and sensitivity of the heat loss responses was significantly lower in the midluteal than in the midfollicular phase, regardless of body site. These effects of the menstrual cycle in group U were not observed in group T. The sweating rate and cutaneous blood flow were significantly higher in group T than in group U, regardless of menstrual phase or body site. Tes threshold for heat loss responses was significantly lower and sensitivity of heat loss responses was significantly greater in group T than in group U in the midluteal phase; however, sensitivity of the sweating response was significantly greater in the midfollicular phase. These results suggest that heat loss responses in group U were inhibited in the midluteal phase compared with in the midfollicular phase. Menstrual cycle had no remarkable effects in group T. Physical training improved heat loss responses, which was more marked in the midluteal than in the midfollicular phase. thermoregulation; estradiol; progesterone; long-term endurance training; heat acclimatization HEAT LOSS RESPONSES, SUCH as sweating and cutaneous vasodilation, during exercise in women differ from those in men because female hormones modify the responses after puberty (3). All studies of the effects of the menstrual cycle on heat loss responses during exercise have demonstrated that the core body temperature thresholds for sweating and cutaneous vasodilation are higher during the luteal phase of the menstrual cycle than during the follicular phase (14,15,17,18,23,31,33,35). In addition, the rise in the core body temperature threshold for sweating and cutaneous vasodilation during the luteal phase is mainly due to elevated progesterone concentrations (8, 31).Conversely, there are conflictin...

The present studies have examined the sequences responsible for regulating transcription of the human DF3 breast carcinoma-associted antigen (MUCI) gene. A region 1656 base pairs upstream to the DF3 transcription initiation site was fused to the chloramphenical acetyltransferase gene. Transient expression assays using a series of deleted constructs demonstrated that the region frown position -618 contains the regulatory sequences necessary for DF3 transcription in human MCF-7 breast cancer cells. Further analysis with internal deletion vectors and heterologous promoter constructs indicated the involvement of cis-acting elements in the fragment extending from positions -598 to -485. By gel retardation and DNA footprinting, we have identified a protein in MCF-7 cells that recognizes sequences between positions -505 and -485. The results of Southwestern studies demonstrate that this protein has an apparent molecular mass of 45 kDa. Taken together, these results suggest that DF3 gene transcription is regulated by a previously undescribed transacting factor.

The murine monoclonal antibody (MAb), designated DF3, reacts with a 300,000-mol wt mammary epithelial antigen. A sequential double-determinant radioimmunoassay (RIA) has been developed to monitor circulating DF3 antigen. Using this assay, we have demonstrated that 33 of 36 normal women had plasma RIA antigen levels < 150 U/mi. In contrast, 33 of 43 patients (76%) with metastatic breast cancer had RIA DF3 antigen levels _ 150 U/ml. The difference between these two groups was statistically significant (P < 0.001). Similar results have been obtained with a double-determinant enzyme-linked immunoassay (EIA). Only 6 of 111 age-matched normal subjects had EIA DF3 antigens levels _ 30 U/ml, while 42 of 58 patients (72%) with breast cancer had levels equal to or above this value. Thus, similar patterns of specificity are obtained with the EIA or RIA. The elevation of circulating DF3 antigen levels in breast cancer patients has been confirmed by transfer blot assays. MAb DF3 reactivity occurred predominantly with circulating antigens of three different molecular weights ranging from 300,000 to -400,000 mol wt. We also demonstrate that patients with both primary and metastatic breast cancer who were free of detectable disease at the time of sampling have DF3 antigen levels that are similar to those obtained from normal subjects. While patients with hepatoma (27%) and ovarian carcinoma (47%) also had elevated circulating DF3 antigen levels, the results suggest that DF3 antigen levels may be useful in distinguishing breast cancer patients from those with esophageal, gastric, colorectal, pancreatic, and lung carcinomas. Furthermore, the results of the RIA, EIA, and transblot analyses demonstrate that the measurement of circulating DF3 antigen levels provides a new and potentially useful marker to follow the clinical course of patients with metastatic breast cancer.

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A B S T R A C T PurposeIn vitro, in vivo, and epidemiologic studies support a role for selenium in reducing the risk of prostate cancer. Our group previously demonstrated a strong interaction between plasma selenium and the manganese superoxide dismutase (SOD2) gene and incident prostate cancer risk. We now hypothesized that SOD2 modifies the association between selenium level and risk of aggressive prostate cancer at diagnosis. Patients and MethodsWe assessed SOD2 variants and plasma selenium in 489 patients with localized/locally advanced prostate cancer from an ongoing retrospective cohort. Cross-sectional associations with aggressive prostate cancer (ie, stage T2b-3, prostate-specific antigen Ͼ 10 ng/mL, or biopsy Gleason score Ն 7) were evaluated using the 2 test, Cochran-Armitage test for trend, and estimations of relative risk (RR) and 95% CIs. Results SOD2genotype alone was not associated with disease aggressiveness, whereas higher versus lower selenium levels were associated with a slightly increased likelihood of presenting with aggressive disease (RR ϭ 1.35; 95% CI, 0.99 to 1.84). There was evidence of an interaction between SOD2 and selenium levels such that among men with the AA genotype, higher selenium levels were associated with a reduced risk of presenting with aggressive disease (RR ϭ 0.60; 95% CI, 0.32 to 1.12), whereas among men with a V allele, higher selenium levels were associated with an increased risk of aggressive disease (for VV or VA men, RR ϭ 1.82; 95% CI, 1.27 to 2.61; P for interaction ϭ .007). ConclusionThese data suggest that the relationship between circulating selenium levels at diagnosis and prognostic risk of prostate cancer is modified by SOD2 genotype and indicate caution against broad use of selenium supplementation for men with prostate cancer.

What’s known on the subject? and What does the study add? Prior studies have identified potential interaction effects between antioxidant nutrients and germline gene variants with regards to prostate cancer risk. In particular, the rs4880 gene variant in SOD2 (or MnSOD) has been linked to several cancers, including prostate, and appears to interact with antioxidant status and cancer risk. We identified additional variants in SOD2 and SOD1 that may affect risk of prostate cancer, or interact with selenium status to affect prostate cancer risk. OBJECTIVE To study the effects of oxidative stress on prostate cancer development as the exact biological mechanisms behind the relationship remain uncertain. We previously reported a statistically significant interaction between circulating selenium levels, variants in the superoxide dismutase 2 gene (SOD2; rs4880), and risk of developing prostate cancer and presenting with aggressive prostate cancer. PATIENTS AND METHODS We genotyped men with localized/regional prostate cancer for 26 loci across eight genes that are central to cellular antioxidant defence: glutathione peroxidase (GPX1, GPX4), peroxisome proliferator‐activated receptor γ coactivator (PPARGC1A, PPARGC1B), SOD1, SOD2, and SOD3, and ‘X‐ray repair complementing defective repair in Chinese hamster cell 1’ (XRCC1). Among 489 men, we examined the relationships between genotypes, circulating selenium levels, and risk of presenting with aggressive prostate cancer at diagnosis, as defined by stage, grade and prostate‐specific antigen (PSA) level (213 aggressive cases). RESULTS Two variants in SOD2 were significantly associated with the risk of aggressive prostate cancer (rs17884057, odds ratio 0.83, 95% confidence interval 0.70–0.99; and rs4816407, 1.27, 1.02–1.57); men with A alleles at rs2842958 in SOD2 had lower plasma selenium levels (median 116 vs 121.8 µg/L, P= 0.03); and the association between plasma selenium levels and risk of aggressive prostate cancer was modified by SOD1 (rs10432782) and SOD2 (rs2758330). CONCLUSION While this study was cross‐sectional and these associations might be due to chance, further research is warranted on the potential important role of antioxidant defence in prostate cancer.