Positive Human Breast Tumors Research Articles

The Angiogenic Secretome in VEGF overexpressing Breast Cancer Xenografts.

The plasticity of cancer cells and the fluidity of the tumor microenvironment continue to present major challenges in the comprehensive understanding of cancer that is essential to design effective treatments. The tumor interstitial fluid (TIF) encompasses the secretome and holds the key to several of the phenotypic characteristics of cancer. Difficulties in sampling this fluid have resulted in limited characterization of its components. Here we have sampled TIF from triple negative and estrogen receptor (ER)-positive human breast tumor xenografts with or without VEGF overexpression. Angiogenesis-related factors were characterized in the TIF and plasma, to understand the relationship between the TIF and plasma secretomes. Clear differences were observed between the TIF and plasma angiogenic secretomes in triple negative MDA-MB-231 breast cancer xenografts compared to ER-positive MCF-7 xenografts with or without VEGF overexpression that provide new insights into TIF components and the role of VEGF in modifying the angiogenic secretome.

Abstract 3486: Interaction of flaxseed oil with trastuzumab in HER2 overexpressing human breast tumors (BT-474)

Abstract BACKGROUND: Trastuzumab (TRAS) is a first line therapy for breast tumors that overexpress the human epidermal growth factor receptor 2 (HER2) but tumors develop resistance within one year of treatment. Flaxseed oil (FO) rich in n-3 fatty acid alpha linolenic acid (ALA) has been shown to reduce HER2 expression and signalling in estrogen receptor positive human breast tumors. Our objective was to determine whether combining dietary FO with TRAS treatment will enhance TRAS effectiveness in reducing tumor growth. METHODS: Ovariectomized mice with established HER2 overxpressing tumors (BT-474) and estrogen pellet implant were fed the basal diet (BD) (control) or treated with TRAS (1 or 2.5mg/kg body weight) and fed either the BD or BD supplemented with 4% FO for 4 weeks. Palpable tumor area was measured weekly. After animal sacrifice, excised tumors were analyzed using immunohistochemistry for cell proliferation (Ki-67) and apoptosis and using Western blot analysis for HER2, MAPK and Akt. RESULTS: Tumors in the control, TRAS1, TRAS2.5, and TRAS1+FO, had palpable tumor areas that significantly increased (p<0.05) by 164%, 104%, 60% and 107%, respectively at the end of the study compared to week 0 while tumors in the FO+TRAS2.5 group did not significantly increase in size. FO+TRAS2.5 treated tumors also had significantly lower (p<0.05) palpable tumor area compared to the TRAS2.5 and other treated tumors at week 4. There was no difference in palpable tumor area between TRAS1 and FO+TRAS1 treated tumors. FO+TRAS2.5 caused a 39% greater increase in apoptosis (p<0.05) and a 61% greater reduction in cell proliferation (p<0.05) compared to TRAS2.5 treatment alone. Western blots support these results and indicate enhanced reductions in HER2, MAPK and Akt expression when TRAS2.5 treatment was combined with dietary FO. CONCLUSIONS: ALA-rich FO enhanced the effectiveness of TRAS at 2.5mg/kg in reducing tumor growth and the mechanisms may involve reduced cell proliferation and increased apoptosis through modulation of HER2 signalling pathways. This is significant as it could lead to a simple, inexpensive complementary treatment for HER2+ patients being treated with TRAS. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3486.

Tamoxifen and the farnesyl transferase inhibitor FTI-277 synergize to inhibit growth in estrogen receptor-positive breast tumor cell lines.

Farnesyl transferase inhibitors (FTIs) serve to specifically inhibit farnesyl isoprenoid lipid modification of proteins. Although originally developed as anti-Ras oncoprotein drugs, it now appears that these compounds function independently of Ras. FTIs have been shown to inhibit transformation by a variety of mechanisms, including apoptosis involving cytochrome c release from mitochondria. Tamoxifen exhibits both anti-estrogenic and estrogenic properties and is widely used as an estrogen antagonist for the treatment of estrogen receptor (ER) positive human breast tumors. Tamoxifen can induce ER-dependent apoptosis in human breast tumor cells by a mechanism involving the Bcl2/mitochondrial arm of the apoptotic machinery. Since tamoxifen and FTIs may stimulate distinct components of the mitochondrial-based apoptotic machinery, we reasoned that their effects might be synergistic. Here we show that anti-estrogens and an FTI (FTI-277) synergize to inhibit cell growth and enhance cell death in ER positive, human breast tumor cell lines. However, the drugs exhibited only additive effects on an ER negative cell line. Analysis of treated ER positive T-47D cells demonstrated that a synergistic increase in apoptosis was induced, as measured by increased caspase 3 activity. Thus, tamoxifen and FTIs may synergize to promote apoptotic cell death in ER positive human breast tumor cells.

Synthesis, Estrogen Receptor Binding, and Tissue Distribution of a New Iodovinylestradiol Derivative (17α,20E)-21-[ 123I]Iodo-11β-Nitrato-19-Norpregna-1,3,5(10),20-Tetraene-3,17-Diol (E-[ 123I]NIVE)

We have synthesized and evaluated E-11β-nitrato-17α-iodovinylestradiol (E-NIVE; E-3c) and its 123I-labelled form, as a new potential radioligand for imaging of estrogen receptor (ER)-positive human breast tumors. E-[ 123I]NIVE was prepared by stereospecific iododestannylation of the E-tri- n-butylstannylvinyl precursor (E-2c), obtained from reaction of 11β-nitrato-estrone (8) with E-tributylstannylvinyllithium. In competitive binding studies, E-NIVE proved to have high binding affinity for both the rat and the human ER (K i 280–730 pM), without significant binding to human sex hormone binding globulin. Distribution studies in normal and mammary tumor-bearing rats showed specific ER-mediated uptake of E-[ 123I]NIVE in the estrogen target tissues, i.e., uterus, ovaries, pituitary, and hypothalamus, but not in the mammary tumors. Selective retention in these target tissues, including tumor tissue, resulted in significant increases over time for the target tissue-to-muscle uptake ratios, but not for the target tissue-to-fat uptake ratios. The tumor-to-fat uptake ratio even appeared constantly below 1. In the primary estrogen target tissues, E-[ 123I]NIVE displayed high specific ER-mediated uptake and retention, which resulted in moderate target-to-nontarget tissue uptake ratios. In contrast, in tumor tissue, E-[ 123I]NIVE uptake appeared to be rather low and not ER-specific. As a consequence, E-[ 123I]NIVE appears to be a less favorable radioligand for ER imaging in breast cancer than the previously studied stereoisomers of 11β-methoxy-17α-[ 123I]iodovinylestradiol (E- and Z-[ 123I]MIVE; [ 123I]E- and [ 123I]Z-3b).

Target tissue uptake selectivity of three fluorine-substituted progestins: Potential imaging agents for receptor-positive breast tumors

We have studied three new fluorine-substituted progestins ( 1–3) as potential imaging agents for progesterone receptor (PgR)-positive human breast tumors. Two of these are fluorine-substituted analogs of the potent progestin R5020 (promegestone), derived from (21S)-hydroxy R 5020 (RU 27987) and (21 R)-hydroxy R 5020 (RU 27988), known metabolites of R 5020, which have affinities for PgR that are 116 and 4%, respectively (relative to R 5020 = 100%). These precursors were protected as their 3,3-dioxolane derivatives and converted to the 21-trifluoromethanesulfonate derivatives. Fluoride ion displacement, followed by acid-catalyzed deprotection, furnished in good yield the epimeric fluoroanalogs, (21 S)- and (21 R)-fluoro R 5020 ( 1 and 2, affinities for PgR, 11 and 45%, respectively). These compounds were also prepared in 18F labeled form by the same route, in 14–32% overall radiochemical yield (decay corrected; synthesis time 90 min; sp. act. 370–1060 Ci/mmol). In tissue distribution studies in estrogen-primed immature rats, uterus-to-muscle ratios were 4.3 at 1 h for the 21S-epimer and 1.1 for the 21 R-epimer, paralleling their relative binding affinities. Considerable metabolic defluorination was observed. The third fluorine-substituted progestin, DU 41165, has a novel retroprogesterone (9β, 10α) structure, substituted with fluorine at C-6; its binding affinity is 145% relative to R 5020, and it was prepared in tritium-labeled form by acetylation of DU 41231, the 17α-hydroxy precursor, with [ 3H]acetic anhydride. In estrogen-primed immature rats, this compound shows uterus-to-muscle ratios of 15 at 1 h, and 18–71 between 2 and 6 h, suggesting that compounds in this retroprogesterone series may be very promising candidates for selective imaging of PgR-positive tissues and tumors.

Variant estrogen receptor mRNA species detected in human breast cancer biopsy samples.

Thirty to 40% of estrogen receptor (ER) positive human breast tumors are resistant to endocrine therapy. To investigate the possibility that abnormal ER proteins may be associated with this endocrine resistant phenotype we have looked for the presence of abnormally sized ER mRNA in human breast tumor biopsies. Poly(A+) enriched RNA was isolated from 46 human breast tumor biopsies and analyzed by Northern blotting and hybridization with the human estrogen receptor OR-8 cDNA. Seventy percent of the tumors contained detectable 6.5 kilobase (kb) ER mRNA. Some tumors also contained smaller sized ER mRNA of approximately 3.8 and 2.4 kb. These variant sized transcripts were only detected in biopsies where the normal 6.5 kb mRNA was present. In some cases the abundance of the variant mRNAs was equal to or greater than the normal ER mRNA. The variant ER mRNAs were not due to nonspecific RNA degradation nor was there any evidence of gross alteration of the ER gene in tumors where variant mRNAs were detected. ER cDNA fragments representing only the E/F domain of the receptor showed little or no hybridization with the variant sized ER mRNAs, while a ER cDNA fragment covering the A/B, C and D domains hybridized to both the variant and normal ER mRNAs. This suggested that the smaller variant ER mRNAs may be missing some or all of the E/F domain which is thought to contain the steroid hormone binding domain of the receptor. It is hypothesized that if these intermediates were translated into viable proteins they may interfere with the normal ER function.

Steroid Δ 4-5β-reductase in human mammary tumors

Steroid Δ 4-5α- and Δ 4-5β-reductase activity was determined in 16 human mammary tumors and 8 DMBA-induced rat mammary tumors using a spectrophotometric assay. Steroid Δ 4-5α-reductase was present in all tumors investigated while Δ 4-5β-reductase was detected in only 6 estrogen receptor negative human breast tumors and absent in all estrogen receptor positive human breast tumors as well as in all rat mammary tumors. Further support for the presence of Δ 4-5β-reductase was established by using a dual-labelling technique consisting of incubating tumor slices with [ 14C] testosterone and adding [ 3H] etiocholanolone, [ 3H] testosterone and [ 3H]-5α-dihydrotestosterone at the end of the reaction. Following extraction and chromic acid oxidation, 4-andro stenedione, 5β-androstanedione and 5α-androstanedione were isolated and purified, and the constancy of the 14C/ 3H ratio was used as proof of 5α-reductase and 5β-reductase. These results were shown to be consistent with the data obtained using the spectrophotometric assay.