Novel Chemical Strategies for Labeling Small Molecule Ligands for Androgen, Progestin, and Peroxisome Proliferator-Activated Receptors for Imaging Prostate and Breast Cancer and the Heart

Summary of Progress The specific aims of this project can be summarized as follows: • Aim 1: Prepare and evaluate radiolabeled ligands for the peroxisome proliferator-activated receptor (PPAR), a new nuclear hormone receptor target for tumor imaging and hormone therapy. • Aim 2: Prepare steroids labeled with a cyclopentadienyl tricarbonyl technetium or rhenium unit. • Aim 3: Prepare and evaluate other organometallic systems of novel design as ligand mimics and halogenated ligands for nuclear hormone receptor-based tumor imaging. As is described in detail below, we made excellent progress on all three of these aims; the highlights of our progress are the following: • we have prepared the first fluorine-18 labeled analogs of ligands for the PPAR receptor and used these in tissue distribution studies in rats • we have developed three new methods for the synthesis of cyclopentadienyltricarbonyl rhenium and technetium (CpRe(CO)3 and CpTc(CO)3) systems and we have adapted these to the synthesis of steroids labeled with these metals, as well as ligands for other receptor systems • we have prepared a number of fluorine-18 labeled steroidal and non-steroidal androgens and measured their tissue distribution in rats • we have prepared iodine and bromine-labeled progestins with high progesterone receptor binding affinity • we have prepared inorganic metal tricarbonyl complexes and steroid receptor ligands in which the metal tricarbonyl unit is an integral part off the ligand core

[18F]Tosyl fluoride as a versatile [18F]fluoride source for the preparation of 18F-labeled radiopharmaceuticals

Positron emission tomography (PET) is an in vivo imaging technology that utilizes positron-emitting radioisotope-labeled compounds as PET radiotracers that are commonly used in clinic and in various research areas, including oncology, cardiology, and neurology. Fluorine-18 is the most widely used PET-radionuclide and commonly produced by proton bombardment o

MMTV-PyMT and derived Met-1 mouse mammary tumor cells as models for studying the role of the androgen receptor in triple-negative breast cancer progression

Triple-negative breast cancer (TNBC) has a faster rate of metastasis compared to other breast cancer subtypes and no effective targeted therapies are currently FDA-approved. Recent data indicate that the androgen receptor (AR) promotes tumor survival and may serve as a potential therapeutic target in TNBC. Studies of AR in disease progression and the systemic effects of anti-androgens have been hindered by the lack of an AR-positive (AR+) immunocompetent preclinical model. In this study we identified the transgenic MMTV-PyMT (mouse mammary tumor virus-polyoma middle tumor antigen) mouse mammary gland carcinoma model of breast cancer and Met-1 cells derived from this model as tools to study the role of AR in breast cancer progression. AR protein expression was examined in late-stage primary tumors and lung metastases from MMTV-PyMT mice as well as in Met-1 cells by immunohistochemistry (IHC). Sensitivity of Met-1 cells to the AR agonist dihydrotestosterone (DHT) and anti-androgen therapy was examined using cell viability, migration/invasion, and anchorage-independent growth assays. Late-stage primary tumors and lung metastases from MMTV-PyMT mice and Met-1 cells expressed abundant nuclear AR protein, while negative for estrogen and progesterone receptors. Met-1 sensitivity to DHT and AR antagonists demonstrated a reliance on AR for survival, and AR antagonists inhibited invasion and anchorage-independent growth. These data suggest that the MMTV-PyMT model and Met-1 cells may serve as valuable tools for mechanistic studies of the role of AR in disease progression and how anti-androgens affect the tumor microenvironment

Association of PET-based estradiol-challenge test for breast cancer progesterone receptors with response to endocrine therapy

Clinical estrogen receptor (ER) testing for breast cancer is limited in predicting response to endocrine therapy (ET). In this phase 2 clinical trial, authors demonstrate that the responsiveness to ET can be predicted by use of PET/CT with 21-[18F]fluorofuranylnorprogesterone (FFNP) to detect the change in tumor progesterone receptor (PgR) levels after a one-day estradiol challenge

17β-Estradiol dysregulates innate immune responses to Pseudomonas aeruginosa respiratory infection and is modulated by estrogen receptor antagonism

ABSTRACT Females have a more severe clinical course than males in terms of several inflammatory lung conditions. Notably, females with cystic fibrosis (CF) suffer worse outcomes, particularly in the setting of Pseudomonas aeruginosa infection. Sex hormones have been implicated in experimental and clinical studies; however, immune mechanisms responsible for this sex-based disparity are unknown and the specific sex hormone target for therapeutic manipulation has not been identified. The objective of this study was to assess mechanisms behind the impact of female sex hormones on host immune responses to P. aeruginosa . We used wild-type and CF mice, which we hormone manipulated, inoculated with P. aeruginosa , and then examined for outcomes and inflammatory responses. Neutrophils isolated from mice and human subjects were tested for responses to P. aeruginosa . We found that female mice inoculated with P. aeruginosa died earlier and showed slower bacterial clearance than males ( P &lt; 0.0001). Ovariectomized females supplemented with 17β-estradiol succumbed to P. aeruginosa challenge earlier than progesterone- or vehicle-supplemented mice ( P = 0.0003). 17β-Estradiol-treated ovariectomized female mice demonstrated increased lung levels of inflammatory cytokines, and when rendered neutropenic the mortality difference was abrogated. Neutrophils treated with 17β-estradiol demonstrated an enhanced oxidative burst but decreased P. aeruginosa killing and earlier cell necrosis. The estrogen receptor (ER) antagonist ICI 182,780 improved survival in female mice infected with P. aeruginosa and restored neutrophil function. We concluded that ER antagonism rescues estrogen-mediated neutrophil dysfunction and improves survival in response to P. aeruginosa . ER-mediated processes may explain the sex-based mortality gap in CF and other inflammatory lung illnesses, and the ER blockade represents a rational therapeutic strategy. </jats:p

Radiosynthesis and evaluation of talazoparib and its derivatives as PARP-1-targeting agents

Poly (ADP-ribose) polymerase-1 (PARP-1) is a critical enzyme in the DNA repair process and the target of several FDA-approved inhibitors. Several of these inhibitors have been radiolabeled for non-invasive imaging of PARP-1 expression or targeted radiotherapy of PARP-1 expressing tumors. In particular, derivatives of olaparib and rucaparib, which have reduced trapping potency by PARP-1 compared to talazoparib, have been radiolabeled for these purposes. Here, we report the first radiosynthesis of

Endogenous DOPA inhibits melanoma through suppression of CHRM1 signaling

Melanoma risk is 30 times higher in people with lightly pigmented skin versus darkly pigmented skin. Using primary human melanocytes representing the full human skin pigment continuum and preclinical melanoma models, we show that cell-intrinsic differences between dark and light melanocytes regulate melanocyte proliferative capacity and susceptibility to malignant transformation, independent of melanin and ultraviolet exposure. These differences result from dihydroxyphenylalanine (DOPA), a melanin precursor synthesized at higher levels in melanocytes from darkly pigmented skin. We used both high-throughput pharmacologic and genetic in vivo CRISPR screens to determine that DOPA limits melanocyte and melanoma cell proliferation by inhibiting the muscarinic acetylcholine receptor M1 (CHRM1) signaling. Pharmacologic CHRM1 antagonism in melanoma leads to depletion of c-Myc and FOXM1, both of which are proliferation drivers associated with aggressive melanoma. In preclinical mouse melanoma models, pharmacologic inhibition of CHRM1 or FOXM1 inhibited tumor growth. CHRM1 and FOXM1 may be new therapeutic targets for melanoma

Long-Term Administration of Conjugated Estrogen and Bazedoxifene Decreased Murine Fecal β-Glucuronidase Activity Without Impacting Overall Microbiome Community

AbstractConjugated estrogens (CE) and Bazedoxifene (BZA) combination is used to alleviate menopause-associated symptoms in women. CE+BZA undergo first-pass-metabolism in the liver and deconjugation by gut microbiome via β-glucuronidase (GUS) enzyme inside the distal gut. To date, the impact of long-term exposure to CE+BZA on the gut microbiome or GUS activity has not been examined. Our study using an ovariectomized mouse model showed that CE+BZA administration did not affect the overall cecal or fecal microbiome community except that it decreased the abundance of Akkermansia, which was identified as a fecal biomarker correlated with weight gain. The fecal GUS activity was reduced significantly and was positively correlated with the abundance of Lactobacillaceae in the fecal microbiome. We further confirmed in Escherichia coli K12 and Lactobacillus gasseri ADH that Tamoxifen-, 4-hydroxy-Tamoxifen- and Estradiol-Glucuronides competed for GUS activity. Our study for the first time demonstrated that long-term estrogen supplementation directly modulated gut microbial GUS activity. Our findings implicate that long-term estrogen supplementation impacts composition of gut microbiota and microbial activity, which affects estrogen metabolism in the gut. Thus, it is possible to manipulate such activity to improve the efficacy and safety of long-term administered estrogens for postmenopausal women or breast cancer patients.</jats:p

Reprogramming of endothelial gene expression by tamoxifen inhibits angiogenesis and ERα-negative tumor growth.

peer reviewedRationale: 17β-estradiol (E2) can directly promote the growth of ERα-negative cancer cells through activation of endothelial ERα in the tumor microenvironment, thereby increasing a normalized tumor angiogenesis. ERα acts as a transcription factor through its nuclear transcriptional AF-1 and AF-2 transactivation functions, but membrane ERα plays also an important role in endothelium. The present study aims to decipher the respective roles of these two pathways in ERα-negative tumor growth. Moreover, we delineate the actions of tamoxifen, a Selective Estrogen Receptor Modulator (SERM) in ERα-negative tumors growth and angiogenesis, since we recently demonstrated that tamoxifen impacts vasculature functions through complex modulation of ERα activity. Methods: ERα-negative B16K1 cancer cells were grafted into immunocompetent mice mutated for ERα-subfunctions and tumor growths were analyzed in these different models in response to E2 and/or tamoxifen treatment. Furthermore, RNA sequencings were analyzed in endothelial cells in response to these different treatments and validated by RT-qPCR and western blot. Results: We demonstrate that both nuclear and membrane ERα actions are required for the pro-tumoral effects of E2, while tamoxifen totally abrogates the E2-induced in vivo tumor growth, through inhibition of angiogenesis but promotion of vessel normalization. RNA sequencing indicates that tamoxifen inhibits the E2-induced genes, but also initiates a specific transcriptional program that especially regulates angiogenic genes and differentially regulates glycolysis, oxidative phosphorylation and inflammatory responses in endothelial cells. Conclusion: These findings provide evidence that tamoxifen specifically inhibits angiogenesis through a reprogramming of endothelial gene expression via regulation of some transcription factors, that could open new promising strategies to manage cancer therapies affecting the tumor microenvironment of ERα-negative tumors

Free Fatty Acids Rewire Cancer Metabolism in Obesity-Associated Breast Cancer via Estrogen Receptor and mTOR Signaling

Obesity is a risk factor for postmenopausal estrogen receptor alpha (ERα)-positive (ER+) breast cancer. Molecular mechanisms underlying factors from plasma that contribute to this risk and how these mechanisms affect ERα signaling have yet to be elucidated. To identify such mechanisms, we performed whole metabolite and protein profiling in plasma samples from women at high risk for breast cancer, which led us to focus on factors that were differentially present in plasma of obese versus nonobese postmenopausal women. These studies, combined with in vitro assays, identified free fatty acids (FFA) as circulating plasma factors that correlated with increased proliferation and aggressiveness in ER+ breast cancer cells. FFAs activated both the ERα and mTOR pathways and rewired metabolism in breast cancer cells. Pathway preferential estrogen-1 (PaPE-1), which targets ERα and mTOR signaling, was able to block changes induced by FFA and was more effective in the presence of FFA. Collectively, these data suggest a role for obesity-associated gene and metabolic rewiring in providing new targetable vulnerabilities for ER+ breast cancer in postmenopausal women. Furthermore, they provide a basis for preclinical and clinical trials where the impact of agents that target ERα and mTOR signaling cross-talk would be tested to prevent ER+ breast cancers in obese postmenopausal women