Epidermal growth factor in NMU-induced mammary tumors in rats

In this work we analyze the hypothesis that tumors induced by ip N-nitroso-N-methylurea injection express EGF-like peptides and EGF receptors which could be involved in the response to hormone manipulation. EGF receptors (EGFR) were determined in the purified membrane fraction of tumors from control and ovariectomized (OVX) animals and no significant differences were found in either maximal binding capacities (Q) or dissociation constants (Kd) between them. Neither did we observe differences between tumors that regressed (HR) or continued growing (HU) after ovariectomy. In order to test the ability of EGFR to trigger a biological response we measured the production of second messengers inositol triphosphates (IP₃) and cAMP levels; we found that EGF increases IP₃ production in a dose-dependent way, while cAMP levels were not affected. In addition, EGF was able to induce in vitro cell proliferation in a concentration-dependent manner when tested in primary cultures of tumor cells by the clonogenic soft agar technique. EGF/TGF- activity was determined by a radioreceptor assay in tumor cytosols from control and OVX rats. Results showed a trend to lower values in tumors from OVX rats, but no differences between HR and HU tumors. A positive correlation was found between EGF/TGF- activity and progesterone receptor maximal binding capacity. When we tested the action of estradiol and EGF added together to primary cultures of tumor cells we found an additive effect on cell proliferation. The study of steady state mRNA levels showed that E₂ increases PgR and c-myc mRNA levels in HR but not in HU tumors. In conclusion, the autocrine loop EGFR-EGF/TGF- present in all tumors is hormonally regulated, possibly by Pg, but is not related to the tumor response to ovariectomy.     

Expression of estrogen receptor-beta in normal mammary and tumor tissues: is it protective in breast carcinogenesis?

Using messenger RNA (mRNA) in situ hybridization, we investigated estrogen receptor- (ER) mRNA levels in normal mammary, benign breast tumor (BBT), breast cancer (BC), and metastatic lymph node tissues to verify the role of ER in BC development and progression. ER expression was significantly decreased in BC and metastatic lymph node tissues compared with normal mammary and BBT tissues (p < 0.01). The intensity and extent of ER mRNA signals were also significantly lower in BC and metastatic lymph node tissues than in the normal mammary and BBT tissues (p < 0.01). An inverse relationship was found between ER mRNA level and both histologic grade (p = 0.091) and progesterone receptor expression (p = 0.052) with marginal significance, but no significant association was noted between ER expression in cancer tissues and the other clinico-pathologic data. The 3-year distant relapse-free survival probability was found to be independent of ER expression. Collectively, ER mRNA decreases in the process of BC development, but seems to be associated with poor differentiation.     

Growth factor involvement in the multihormonal regulation of MCF-7 breast cancer cell growth in soft agar

Summary The hormone dependency of the MCF-7 breast cancer cell line, while extensively tested in liquid culture, has not been previously evaluated under conditions of anchorage-independent growth in serum-free media. Using the soft agar clonogenic assay, we demonstrate that physiologically relevant concentrations of estradiol (E₂), progesterone (Pg), and prolactin (PRL) similarly stimulated MCF-7 cell colony formation in the absence of serum. Addition of an anti-insulin-like growth factor-I (IGF-I) antibody inhibited E₂- and Pg-stimulated growth, while PRL action was not affected. Similar results were obtained with an anti-IGF-I receptor antibody, except that its inhibitory effect on Pg-induced colony formation was modest and not statistically significant. Administration of either an anti-transforming growth factor- (TGF-) antibody or an anti-epidermal growth factor (EGF) receptor antibody similarly inhibited E₂-stimulated MCF-7 cell growth in soft agar, while neither antibody influenced Pg or PRL effects. Addition of TGF-₁, -₂, -₃ similarly suppressed MCF-7 cell colony formation in a dose dependent manner to a degree comparable to that observed with 4-OH-tamoxifen (4-OH-T). Furthermore, the growth inhibitory effect of 4-OH-T was completely reversed by an anti-TGF- antibody. We conclude that IGFs and TGF- are important mediators of E₂-stimulated MCF-7 cell growth in soft agar. IGFs may also be playing a role in Pg action, while neither growth factor is involved in PRL-stimulated colony formation. Finally, TGF- appears to be an important mediator of antiestrogen-induced inhibition of tumor growth.Abbreviations IGF insulin-like growth factor - TGF transforming growth factor - E₂ estradiol-17 - Pg progesterone - PRL prolactin - 4-OH-T 4-hydroxy-tamoxifen - IMEM improved minimal essential medium     

Sex hormone-induced mammary carcinogenesis in female Noble rats: detection of differentially expressed genes

Breast cancer is the most common cancer and the second most frequent cause of cancer death in women. Epidemiological data has recognized that an increased cumulative exposure to estrogen is the common tie linking most of the established risk factors for breast cancer. Sex hormone-induced mammary gland carcinogenesis of the Noble rat (using testosterone and 17-estradiol) resembles that of the human counterpart in its growth pattern as well as the histopathology of the tumors induced. This model may provide a paradigm for examination of genetic alterations and changes in gene expression between different histological groups and to make inferences about the role of known and putative oncogenes and tumor suppressor genes. We studied the gene expression profile during sex hormone-induced mammary carcinogenesis using a cDNA array technique; the results were further confirmed by RT-PCR, western blotting and immunohistochemical analyses. From the 10 differentially expressed genes identified, we have studied four highly overexpressed genes, two cell cycle/growth control regulators, the cyclins D1 and D2, a growth factor, IGF-2 and a cytokine TNF-. Cyclins D1 and D2 were highly expressed in the nuclei of carcinoma cells but at low levels in the nuclei of the hyperplastic and normal mammary tissue. IGF-2 was found to expressed in the cytoplasm of the carcinoma cells but not in the stromal cells. Western blot showed expression of big IGF-2 consistent with the tumor derived truncated forms of pro-IGF-2. The matured circulating IGF-2 at 7.5 kDa identified in the serum was not expressed in any of the breast tissue samples. TNF- expression was found not only in the macrophages but also in the mammary carcinoma cells. The result of the present study provides some information on the molecular basis of this sex hormone-induced mammary carcinogenesis and the role of these proteins in tumor progression.     

The anti-proliferative effect of suramin towards tamoxifen-sensitive and resistant human breast cancer cell lines in relation to expression of receptors for epidermal growth factor and insulin-like growth factor-I: Growth stimulation in the presence of tamoxifen

Background: A significant proportion of breast cancer patients receiving tamoxifen therapy relapse during treatment following acquisition of tamoxifen-resistant or oestrogen-independent phenotypes. The mechanism behind this rapid progression to oestrogen autonomy is at present unclear and further treatment modalities are limited. Suramin represents a novel potential second line therapy. The mechanism of the antineoplastic activity of suramin is not completely understood, although the drug binds to many growth factors including epidermal growth factor and insulin-like growth factors and can also dissociate growth factors from their receptors. In this study we have related suramin sensitivity to the expression of receptors for epidermal growth factor and insulin-like growth factor-I in a number of breast cancer cell lines including lines resistant to tamoxifen.Materials and methods: The anti-proliferative effects of suramin were investigated in two oestrogen dependent breast cancer lines (ZR-75-1 and MCF-7), oestrogen independent (ZR-PR-LT) and tamoxifen resistant (ZR-75-9a1) variants of ZR-75-1 and a tamoxifen resistant (LY2) variant of MCF-7. Full dose response curves were constructed and IC⁵⁰values determined for each cell line. Sensitivity to suramin was correlated with the level of expressio n of receptors for epidermal growth factor (EGFR) and insulin-like growth factor-I (IGFR). On observing stimulation of cell proliferation by suramin in the tamoxifen resistant cell lines in the presence of tamoxifen we also investigated the possible role of suramin sequestration of transforming growth factor- in mediating this effect.Results: All cell lines exhibited a dose- and time-dependent response to suramin treatment. Tamoxifen resistant ZR-75-9a1 cells (day 6 IC⁵⁰85 µg ml^–1) were more resistant to suramin than oestrogen independent ZR-PR-LT cells (day 6 IC⁵⁰45 µg ml^–1), and the parent ZR-75-1 line (day 6 IC⁵⁰56 µg ml^–1). Increased sensitivity to suramin was associated with increased expression of IGFR and decreased expression of EGFR. Tamoxifen resistant LY2 cells were significantly more sensitive to suramin (day 6 IC⁵⁰70 µg ml^–1) than MCF-7 cells (day 6 IC⁵⁰350 µg ml^–1). Both IGFR and EGFR expression by LY2 cells was lower than in the parent line. The antioestrogen-resistant ZR-75-9a1 and LY2 lines grown in the presence of 8 µM tamoxifen were growth stimulated by concentrations of the drug below 100 µg/ml. As growth stimulation observed in the presence of tamoxifen may have been due to suramin sequestration of tamoxifen induced TGF-1 secretion we also investigated the response of the cells to this peptide in the presence and absence of suramin. All cell lines were growth inhibited by TGF-1 except ZR-75-9a1 which was unresponsive. Responses to TGF-1 were modified in the presence of 100 µg suramin ml^–1 although TGF-1 was unable to mimic the ability of tamoxifen to stimulate proliferation in the presence of suramin.Conclusions: These results suggest that for ZR-75-1 cells and variants, increased sensitivity to suramin is associated with an increase in expression of IGFR and a decrease in EGFR numbers. However, tamoxifen resistant LY2 cells, in which both IGFR and EGFR expression is reduced were considerably more sensitive than parental MCF-7 cells suggesting that there is no clear relationship between EGFR and IGFR expression and suramin sensitivity. The unexpected stimulation of cell proliferation of the tamoxifen resistant variants by suramin in the presence of tamoxifen could not be explained by suramin sequestration of transforming growth factor- and the mechanism of this interaction remains unclear.     

Immunological detection and quantitation of alpha transforming growth factors in human breast carcinoma cells

Summary Alpha transforming growth factors (TGFs) were immunologically detected in the concentrated conditioned medium (CM) prepared from four human breast cancer cell lines and from primary cultures of human mammary epithelial cells, and in the tissue extracts prepared from normal, benign, and malignant breast biopsies. Immunoreactive TGFs were quantitated by a competitive radioimmunoassay (RIA) using affinity-purified polyclonal sheep anti-rat TGF antibodies which react with human TGF but not with human epidermal growth factor (EGF). The relative level of RIA-detectable TGFs in the CM from the breast cancer cell lines MCF-7, ZR-75-1, T47-D, and MDA-MB-231, and from the CM of primary cultures of human mammary epithelial cells, ranged from 0.02 to 0.85 ng/ml. MCF-7 or ZR-75-1 cells grown in the presence of 17-estradiol (10^–8 M) for 48 h were found to release two- to three-fold more TGFs into their CM than the same cells grown in the absence of estrogen. In detergent extracts prepared from normal breast tissue, a benign fibrocystic lesion, fibroadenomas and primary breast carcinomas, the relative TGF concentrations were found to range from 1.5 to 6 ng/mg cell protein. No significant correlations were found between the TGF levels and the pathological state of the tissues, the estrogen receptor status of the tumors, or the relative amounts of theras gene protein p21ras in the tissues as determined by Western immunoblot analysis. The question of biological relevancy of TGF for human mammary tumors will require further studies on (a) synthesis and turnover of TGF, (b) the relationship between immunoreactivity and biological activity of TGF, and (c) differences in biological responsiveness of mammary tumor cells.     

Verotoxins inhibit the growth of and induce apoptosis in human astrocytoma cells

Verotoxin 1 (VT1) is an E. coli toxin comprising an A subunit with N-glycanase activity, and five smaller B subunits capable of binding to the functional receptor globotriaosylceramide (Gal1-4-Gal1-4-Glcceramide-Gb3). VT is implicated in hemorrhagic colitis and the more serious hemolytic uremic syndrome. VT1 is active against various tumor cell lines in vitro and in vivo. To extend the anti-cancer spectrum of activity of VT to human brain tumors, in the present analysis we studied the effects of VT on the growth of 6 permanent human astrocytoma cell lines. All astrocytoma cell lines analyzed express Gb3 and were sensitive to VT-1 at a dose of 50 ng/ml, but sensitivity was not proportional to the relative Gb3 concentration. VT induced apoptosis in these cells was shown by electron microscopy. Morphological evidence (nuclear shrinkage and chromatin condensation) of apoptosis could be clearly distinguished 1.5 hrs after toxin addition. Ultrastructural preservation of organelles was observed in conjunction with blebbing of the plasma membrane, condensation of chromatin within the nucleus and nuclear shrinkage. Apoptosis was also induced by the recombinant toxin B subunit alone, suggesting that the ligation of Gb3 is the primary induction mechanism. These studies indicate that verotoxin/Gb3 targetting may provide a novel basis for the inhibition of astrocytoma tumour cell growth.     

Quantification of thrombospondin-1 secretion and expression of αvβ3 and α3β1 integrins and syndecan-1 as cell-surface receptors for thrombospondin-1 in malignant glioma cells

Malignant glioma cells secrete thrombospondin-1 (TSP-1) which participates in the motility of glioma cells, and binds to cell surface v3 and 31 integrins, and syndecan-1. This study evaluated the amount of TSP-1 secretion from malignant glioma cells, and the expression of v3 and 31 integrins, and syndecan-1. The amounts of TSP-1 in the supernatants from 10 malignant glioma cell lines and eight non-glioma malignant tumor cell lines were measured by enzyme-linked immunosorbent assay. Expression of v3 and 31 integrins, and syndecan-1 were examined by flow cytometry. The amounts of TSP-1 secreted by malignant glioma cells were 43 to 2431 ng/1 × 10⁶ cells/24 h (mean ± SD=626 ± 792). Seven of 10 glioma cell lines secreted more than 100 ng of TSP-1 and three of these cell lines secreted more than 1 g. Seven of eight non-glioma cell lines secreted less than 100 0ng of TSP-1. All glioma cell lines expressed 31 integrin and syndecan-1, and seven of 10 glioma cell lines expressed v3 integrin. Treatment of the glioma cell lines with TGF-2 did not change the expression of v3 integrin. These results suggest that malignant glioma cells secrete high levels of TSP-1, which may be important in the migration of glioma cells via interactions with v3 and 31 integrins, and syndecan-1.     

Interferon effect on glycosaminoglycans in mouse gliomain vitro

Summary The effect of mouse interferon / (MuIFN /) on the production of glycosaminoglycans (GAGs) by mouse glioma G-26in vitro was evaluated. Two GAG species secreted extracellularly by the mouse glioma G-26 were isolated using cellulose acetate electrophoresis. They were identified as hyaluronic acid (HA) and chondroitin sulfate (CS) following enzymatic digestion with enzymes: hyaluronidase and chondroitinase ABC. Further characterization of CS by enzymatic digestion with specific chondroitinases for chondroitin 4-sulfate (CSA) and chondroitin 6-sulfate (CSC), revealed that the isolated CS was neither CSA nor CSC. Therefore, it may be either chondroitin sulfate B (CSB) (dermatan sulfate) or one of the chontroitin sulfate isomers (D-H).The three day incubation of glioma G-26 cells with 8×10-8×104 U/ml of MuIFN / resulted in a dose dependent inhibition of cell proliferation measured by³H-thymidine incorporation and the MTT assay. The significant decrease of the CS (p < 0.008) but not the HA level, (measured densitometrically), was observed following 72 hours (hrs) incubation of G-26 cells with 8 × 10³ U/ml of MuIFN / (IFN treated cells: 0.03 ± 0.007 integrated optical density (IOD); control cells: 0.07 ± 0.01 IOD). The decreased CS production may be the underlying cause of IFN mediated inhibition of glioma cell proliferation.     

Candidate metastasis-associated genes of the rat 13762NF mammary adenocarcinoma

Summary Differential hybridization was used to isolate genes potentially involved in the process of metastasis. Ten complementary DNAs (cDNAs) that were differentially expressed between a highly metastatic (MTLn3) and a nonmetastatic (MTC.4) line of the rat 13762NF mammary adenocarcinoma were isolated and sequenced. Examination of the EMBL/GenBank database revealed that one of the genes had a high degree of homology (98.8%) to annexin I (also known as calpactin II). Quantitative analysis of Northern blot hybridizations showed that the annexin I-like sequence was expressed 4- to 7-fold higher in MTLn3 than in MTC.4 cells. Steady state mRNA levels were also low in MTLn2, a cell line of low metastatic potential closely related to MTLn3, but were not related to metastatic potential in colon adenocarcinoma or melanoma cells. Two of the cDNAs (designated 8.11 and 10.14) were found to be novel. The expression of 10.14 mRNA (3.2 kb) was 4-fold higher in MTLn3 than in MTC.4 cells. Sequencing of the 10.14 cDNA (2.2 kb) revealed a putative open reading frame of 583 amino acids that was also novel. Expression of 8.11 mRNA (>7 kb) inversely correlated with metastatic potential. Another differentially transcribed gene was highly homologous to ERK2 (extracellular signal related kinase 2), a mitogen-activated protein kinase (MAPK). Northern analysis of ERK2 expression revealed 3-fold higher amounts of a 1.3 kb mRNA in MTLn3 than in MTC.4 cells. Higher levels of ERK2 mRNA were generally seen in the more metastatic human colon but not in melanoma cell lines. We also corroborated the work of Taniguchi (Nucl Acids Res 19:6949, 1991) by independently identifying EF-1 as a putative metastasis-associated gene.Supported by an educational grant from CIBA-GEIGY Corp., at the San Antonio Breast Cancer Symposium, Dec. 8, 1992.     

Urinary 2-hydroxyestrone/16alpha-hydroxyestrone ratio and family history of breast cancer in premenopausal women

It has been hypothesized that the ratio of urinary 2-hydroxyestrone to 16-hydroxyestrone (2-OHE₁/16-OHE₁ represents a biomarker for breast cancer risk; the lower the ratio the higher the risk. We obtained early morning urine samples from 70 'high risk' premenopausal women who had a first degree family history of breast cancer and 27 'low risk' women with no such history. Five estrogen metabolites in urine were determined: 2-OHE₁, 16-OHE₁, estrone (E₁), estradiol (E₂), and estriol (E₃) conjugates. We compared geometric mean levels of each metabolite adjusted for age and weight. 'High risk' women did not have elevated levels of any of these metabolites. Instead, we observed decrements of 3–27% in women with a family history of breast cancer compared with women without such history, this difference was statistically significant for E₂, 2- OHE₁, and 16-OHE₁. The ratio of 2-OHE₁/16-OHE₁ was identical in women with and without a family history of breast cancer. These results were unchanged, when additionally adjusted for recent intake of alcohol or cruciferous vegetables. Our data suggest that among premenopausal women, family history is not associated with higher urinary estrogen levels or a lower ratio of urinary 2-OHE₁/16-OHE₁.     

Manganese superoxide dismutase (MnSOD) polymorphism,alpha-tocopherol supplementation and prostate cancer risk in the alpha-tocopherol,beta-carotene cancer prevention study (Finland)

Objective: Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme that plays a key role in protecting the cell from oxidative damage. A polymorphism in the mitochondrial targeting sequence (a valine to alanine substitution), thought to alter transport of the enzyme into mitochondria, has been associated with increased risk for breast cancer with a more pronounced association among women with low intake of dietary antioxidants. We examined the role of MnSOD in the development of prostate cancer in a large, randomized cancer prevention trial of male smokers, the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study. We hypothesized that MnSOD may be associated with prostate cancer and that long-term antioxidant supplementation (-tocopherol 50 mg/day for five to eight years) could modify the effect on risk. Methods: Logistic regression was used to estimate these associations among 197 cases and 190 controls genotyped and matched for age, intervention group, and clinic. Results: Men homozygous for the MnSOD ala allele had a 70% increase in risk over men homozygous for the val allele (odds ratio, OR = 1.72, 95% confidence interval, CI = 0.96–3.08, p = 0.07). Supplementation with -tocopherol had no impact on the MnSOD–prostate cancer association. Although there was no difference in the association with disease stage, men homozygous for MnSOD ala (compared to MnSOD val/val or val/ala) showed a three-fold risk increase for high-grade tumors (OR = 2.72, 95% CI: 1.15–6.40, p = 0.02). Conclusion: These data suggest an effect of the MnSOD ala/ala genotype on the development of prostate cancer. Our observation of a stronger association with high-grade tumors may have prognostic implications that should also be pursued.     

Epidermal growth factor-related peptides in the pathogenesis of human breast cancer

Summary A number of different epidermal growth factor (EGF)-related peptides such as EGF, transforming growth factor (TGF), amphiregulin (AR), heregulin (HRG), and cripto-1 (CR-1), are coexpressed to varying degrees in both normal and malignant mammary epithelial cells. However, in general the frequency and level of expression of TGF, AR, and CR-1 are higher in malignant breast epithelial cells than in normal mammary epithelium. In addition, several of these peptides such as TGF and AR can function as autocrine and/or juxtacrine growth factors in mammary epithelial cells, and their expression is stringently regulated by mammotrophic hormones such as estrogens, activated proto-oncogenes that have been implicated in the pathogenesis of breast cancer, and other growth factors. The redundancy of expression that is observed for a number of these structurally related peptides in both normal and malignant mammary epithelial cells suggests that some of these peptides may be involved in regulating other aspects of cellular behavior such as differentiation in addition to proliferation.     

Dietary n – 6 and n – 3 Polyunsaturated Fatty Acids and Prostate Cancer Risk: A Review of Epidemiological and Experimental Evidence

OBJECTIVE: This study reviews epidemiological and experimental works dealing with the effects of dietary n -6 or n -3 polyunsaturated fatty acids (PUFA) on prostate cancer (PCa) development and PCa risk. METHODS: Systematic literature searches were made using Medline. The epidemiological studies reviewed (ecological, case-control, cohorts, and nested case-control) were those having tested the association of PCa risk with the dietary intake or the blood or adipose tissue levels of PUFA ( n -6 PUFA, n -3 PUFA, long-chain n -3 PUFA, linoleic acid, alpha -linolenic acid, arachidonic acid, eicosapentaenoic acid, docosahexaenoic acid), and with the dietary intake of fish and seafood. Experimental studies dealing with the effects of PUFA on PCa development in animal models or with PCa cell growth in vitro were also reviewed, as well as studies on the mechanisms of the effects of PUFA on PCa. RESULTS: There is no or little evidence of an association of linoleic or arachidonic acids with PCa risk. Most epidemiological studies failed to find an association of PCa risk with fish or long-chain n -3 PUFA intake, but two recent cohort studies did find an inverse association of fish consumption with the risk of the latest stages of PCa. alpha -linolenic acid intake was associated with an increase of PCa risk in a majority of epidemiological studies, but other studies did not find this association. Experimental work in vitro and in vivo, as well as mechanistic studies, support a protective effect of long-chain n -3 PUFA on PCa, but data on the effects of linoleic and alpha -linolenic acids are scarce. CONCLUSIONS: Long-chain n -3 PUFA from fish are possible promising nutrients for the dietary prevention of PCa, but to-date with little epidemiological support. In contrast, studies suggest that alpha -linolenic acid intake might be a risk factor. New work, both epidemiological and experimental, is awaited to clarify these results.     

Prognostic relevance of epidermal growth factor receptor (EGF-R) and c-neu/erbB2 expression in glioblastomas (GBMs)

Summary Seventeen untreated primary adult glioblastomas were analyzed using immunocytochemistry for the expression of EGF-R, c-neu/erbB2, TGF-, and phosphotyrosine. Patients were divided by median survival into long-term or short-term survivors (LTS, N=10, median > 4 years; versus STS, N=7, median 61 weeks). There were no significant differences between the two groups in terms of age, extent of resection, post-operative Karnofsky status, or treatment. Diagnostic sections from each tumor were stained with antibodies to EGF-R, c-neu/erbB2, TGF- and phosphotyrosine. Double-labelling for TGF- and EGF-R was also performed. All 10/10 LTS were considered to be EGF-R negative/scant, while 4/7 STS were EGF-R positive. EGF-R negativity significantly correlated with long-term survival. The differences in c-neu/erbB2 expression did not reach significance. However, 4/7 STS were positive for both proteins and 76% of the 17 cases were either double negative or positive for EGF-R and c-neu/erbB2. TGF- and phosphotyrosine were frequently expressed, but neither were prognostic. Recurrent tumors were studied in 7 STS. EGF-R expression was increased in 4/7 of these cases and c-neu/erbB2 was increased in all 7 cases, compared to the pretreatment baselines. Increased expression of these proteins in glioblastomas may be associated with aggressive clinical behavior and treatment resistance.     

1α-hydroxyvitamin D3, hypercalcemia,and growth suppression of 7,12-dimethylbenz[a]anthracene-induced rat mammary tumors

Summary 1-hydroxyvitamin D₃ [1(OH)D₃] was administered to female Sprague-Dawley rats with 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors. 1(OH)D₃ suppressed the growth of the rat mammary tumors dose-dependently, and in the high dose groups treated with 0.5–1.0µg/kg of 1(OH)D₃, significant inhibition of tumor growth was observed. But daily oral administration of 1(OH)D₃ for four consecutive weeks caused side effects such as hypercalcemia and weight loss. We compared 0.5 µg/kg of 1(OH)D₃ three times weekly with the same dose six times weekly to discover whether or not the side effects can be reduced by treatment schedule. Both groups showed a significant oncostatic effect, compared with the control group, while the side effects were relieved in the three times weekly group. Regarding estrogen receptors (ER) in the tumors, there was no significant difference among the groups. These results suggested that the antitumor effect of 1(OH)D₃ on DMBA-induced mammary tumors was not related to ER status. Combined use of 1(OH)D₃ with 5-fluorouracil (5-FU) or medroxyprogesterone acetate (MPA) was also examined. No significant augmentation of the antitumor effect was seen in the two combinations, although the combined therapy with MPA showed a significant inhibition of weight loss in the rats.     

A 150-kDa glycoprotein isolated from Solanum nigrum L. has cytotoxic and apoptotic effects by inhibiting the effects of protein kinase C alpha, nuclear factor-kappa B and inducible nitric oxide in HCT-116 cells

This study was carried out to investigate the anticancer effects of a 150-kDa glycoprotein isolated from Solanum nigrum L. (SNL glycoprotein) on spontaneously and experimentally induced tumor promotion in HCT-116 cells. For spontaneously induced tumor promotion, we evaluated the cytotoxic and apoptotic effects in HCT-116 cells using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT), DNA fragmentation, and H33342 and ethidium bromide staining assays. SNL glycoprotein had remarkable, dose-dependent cytotoxic and apoptosis-inducing effects at low concentrations. For experimentally induced tumor promotion, we investigated whether the SNL glycoprotein was able to regulate the activity of protein kinase C alpha (PKC), the DNA binding activation of nuclear factor-kappa B (NF-B), the activity of NF-B protein, and the production of nitric oxide (NO) in HCT-116 cells stimulated with 12-O-tetradecanoylphorbol 13-acetate (TPA) using electrophoretic mobility shift assays (EMSA), Western blot analysis, and NO assays. As expected, SNL glycoprotein dose-dependently inhibited PKC translocation, NF-B DNA binding activity, NF-B protein activity and NO production in HCT-116 cells stimulated with TPA (61.68 ng/ml, 100 nM). Collectively, these results suggest that SNL glycoprotein can induce apoptosis through the modulation of signal mediators. Therefore, we speculate that it could be used as a chemotherapy agent even at low concentrations in HCT-116 cells.     

Vitamin E concentration in breast adipose tissue of breast cancer patients (Kuopio, Finland)

Previous data on animals and humans suggest that vitamin E may be a protective factor against cancer. A low dietary vitamin E intake has been suggested to increase the risk of breast cancer. We examined the dietary intake and the concentration of vitamin E in breast adipose tissue of women in Kuopio, Finland, diagnosed between 1990 and 1992 with benign breast disease (n=34) and with breast cancer (n=32). In postmenopausal women, lower dietary intake (P=0.006) and a smaller concentration of vitamin E in breast adipose tissue (P=0.024) were observed in breast cancer patients than in subjects with benign breast disease. Partial correlation showed that the vitamin E concentration in the breast adipose tissue correlated positively with the dietary intake of vitamin E (r=0.25, P=0.023), indicating that the vitamin E concentration in breast adipose tissue reflects the dietary intake of vitamin E.Drs Zhu and Uusitupa are with the Department of Clinical Nutrition, University of Kuopio, Finland, Dr Parviainen is with the Laboratory of Helsinki University, Central Hospital, Helsinki, Finland. Drs Männistö and Pietinen are with the Department of Nutrition, National Public Health Institute, Helsinki, Finland Dr Eskelinen is with the Department of Surgery, and Dr Syrjänen is with the Department of Pathology, University of Kuopio. Address correspondence to Professor Uusitupa, Department of Clinical Nutrition, University of Kuopio, P.O. Box 1627, FIN-70211 Kuopio, Finland. This study was supported by research grants from the Finnish Cancer Society, and by EVO funding for the Breast Cancer Project of Kuopio Cancer Research Center from the Kuopio University Hospital, Finland.     

Structure-activity relations in carcinogenesis by N-nitroso compounds

For a large number of N-nitroso compounds a comparison of their carcinogenic effects in rats and Syrian golden hamsters has been made. Nitrosamines, which require metabolic activation, and nitrosoalkylamides, which do not, produce quite different tumor responses. There are also large differences in the types of tumor induced in rats and in hamsters. In all the studies doses of the various compounds, equimolar to the extent that was possible, are administered orally. Continuous doses (in drinking water or food) often produce a response different from that after administration of the same compound in pulsed doses (by gavage), even though the same total dose is delivered. Continuous doses of nitrosamines are usually more effective than pulsed doses, but with the nitrosoalkylureas, the reverse is more generally the case. Rat and hamster liver is a common target of many nitrosamines, but rarely of nitrosamides. The most common site of tumor induction in rats by N-nitroso compounds is the esophagus, but the hamster esophagus never responds. The pancreas duct of the hamster is a common target of nitrosamines containing a -oxygenated propyl group, but pancreas duct tumors are never seen in rats. Nitrosomethyl-n-alkylamines (with an even numbered carbon chain) induce bladder tumors in rats and hamsters. Many nitrosoalkylureas induce tumors of the nervous system in rats, as well as a great variety of other tumors. In hamsters, nitrosoalkylureas give rise only to tumors of the forestomach and spleen, but no tumors of the nervous system. The similar carcinogenic actions of certain groups of N-nitroso compounds can be related to their generation, directly or by metabolism, of similar simple moieties having certain organs as their target. Note. In this article all reference citations are enclosed in brackets, while the N-nitroso compounds referred to in Table 1 are enclosed in parentheses and italicized.