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Estrogens in Prostate Cancer
Posted December 3, 2007
Shuk-Mei Ho, Ph.D., University of Cincinnati, Cincinnati, Ohio

Dr. Shuk-Mei Ho of the University of Cincinnati has received multiple awards from the Department of Defense Prostate Cancer Research Program for Fiscal Years 1997, 2000, 2003, and 2005 to examine the involvement of the female hormone estrogen and estrogen-like compounds in prostate cancer. Her earlier funded work identified paradoxical roles for estrogen receptor (ER) beta in inhibiting prostate cancer progression but promoting metastasis. More recently, in conjunction with Dr. Gail Prins at the University of Illinois at Chicago, she demonstrated for the first time that increased prenatal exposure to environmentally relevant doses of agents that mimic the actions of estrogen predisposes prostate tissue to precancerous lesions and tumor development later in life. One such agent, bisphenol A (BPA), is commonly found in plastics and also occurs naturally in placental and fetal tissue. Dr. Ho's group has also identified a gene that is epigenetically regulated by estradiol or BPA exposure. This gene is a critical link in establishing a mechanism of xenoestrogen carcinogenesis. The researcher's findings exposing the critical role of estrogens and ER beta in prostate cancer have increased interest in developing new preventive or treatment agents that block harmful estrogenic signaling in the prostate. Dr. Ho's discovery of multiple variants (isoforms) of ER beta with different tissue distributions and actions should facilitate the development of agents that specifically inhibit the deleterious effects of estrogen in prostatic tissue with minimal side effects.

Publications:

Leung YK, Mak P, Hassan S, Ho SM. 2006. Estrogen receptor (ER)-beta isoforms: A key to understanding ER-beta signaling. Proceedings of the National Academy of Sciences U S A 103:13162-13167. Erratum: Proceedings of the National Academy of Sciences U S A 103:14977.

Ho SM, Tang WY, Belmonte de Frausto J, Prins GS. 2006. Developmental exposure to estradiol and bisphenol A increases susceptibility to prostate carcinogenesis and epigenetically regulates phosphodiesterase type 4 variant 4. Cancer Research 66:5624-5632.

Ho SM, Leung YK, and Chung I. 2006. Estrogens and antiestrogens as etiological factors and therapeutics for prostate cancer. Annals of the New York Academy of Sciences 1089:177-193.

Zhu X, Leav I, Leung YK, Wu M, Liu Q, Gao Y, McNeal JE, and Ho SM. 2004. Dynamic regulation of estrogen receptor-beta expression by DNA methylation during prostate cancer development and metastasis. American Journal of Pathology 164:2003-2012.

Leav I, Lau KM, Adams JY, McNeal JE, Taplin ME, Wang J, Singh H, and Ho SM. 2001. Comparative studies of the estrogen receptors beta and alpha and the androgen receptor in normal human prostate glands, dysplasia, and in primary and metastatic carcinoma. American Journal of Pathology 159:79-92.

Lau KM, LaSpina M, Long J, Ho SM. 2000. Expression of estrogen receptor (ER)-alpha and ER-beta in normal and malignant prostatic epithelial cells: regulation by methylation and involvement in growth regulation. Cancer Research 60:3175-3182.

Link:

Abstract: Estrogen Receptor-beta Hypermethylation and Prostate Carcinogenesis

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Magnetic Resonance Spectroscopy: An Objective Technique for the Quantification of Prostate Cancer Pathologies
Posted November 20, 2007
Leo Cheng, Ph.D., Massachusetts General Hospital, Boston, Massachusetts

In the past, high-resolution magic-angle-spinning (HRMAS) spectroscopy was restricted to the determination of the physical structures of small molecules and some proteins. Recently, however, this branch of magnetic resonance spectroscopy has been extended to studies of more complex molecules in cells and tissues isolated from patients. Dr. Leo Cheng of Massachusetts General Hospital, recipient of a Fiscal Year 2003 Department of Defense Prostate Cancer Research Program Idea Development Award, has taken full advantage of this new technique and has begun to translate this imaging modality into the clinic for prostate cancer detection and diagnosis. By definition, HRMAS spectroscopy requires that samples be spun at very high rates, which can disrupt tissue architecture and prevent accurate pathological analysis. To address this roadblock, Dr. Cheng engineered a method that provides quantifiable, high-resolution data for prostate metabolites at spinning speeds that do not damage prostate tissue architecture. Dr. Cheng focused on three lipid molecules, choline, phosphocholine (PC), and glycerophosphocholine (GPC), whose metabolism often increases with the progression of prostate cancer. In order to measure these metabolites, Dr. Cheng has integrated 31P-edited 1H spectroscopy with HRMAS techniques to reliably quantify the concentrations of choline, PC, and GPC in human prostate biological samples based on the unique phosphorus signal of each metabolite. The metabolic profile generated by these molecules was remarkably consistent, even after long-term storage. When combined, HRMAS and 31P-edited 1H spectroscopy may one day measure the aberrant metabolism of a tumor for the diagnosis and treatment of prostate cancer in the clinic.

Publications:

Taylor JL, Wu CL, Cory D, Gonzalez RG, Bielecki A, Cheng LL. 2003. High-resolution magic angle spinning proton NMR analysis of human prostate tissue with slow spinning rates. Magn Reson Med 50:627-632.

Wu CL, Taylor JL, He WL, Zepeda AG, Halpern EF, Bielecki A, Gonzalez RG, Cheng LL. 2003. Proton high resolution magic angle spinning nmr analysis of fresh and previously frozen tissue of human prostate. Magn Reson Med 50:1307-1311.

Burns MA, He W, Wu CL, Cheng LL. 2004. Quantitative pathology in tissue MR spectroscopy based human prostate metabolomics. Technol Cancer Res Treat 3: 591-598.

Cheng LL, Burns MA, Taylor JL, He WL, Halpern EF, McDougal WS, Wu, CL. 2005. Metabolic characterization of human prostate cancer with tissue magnetic resonance spectroscopy. Cancer Res 65:3030-3034

Link:

Abstract: Magnetic Resonance Spectroscopy: An Objective Technique for the Quantification of Prostate Cancer Pathologies

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Regulation of Androgen Receptor Function by ErbB Receptor Tyrosine Kinases
Posted November 9, 2007
Ingo Mellinghoff, M.D., University of California, Los Angeles, California

While androgen deprivation therapy is initially very successful in reducing the tumor burden associated with early stage prostate cancer, the adaptability of the disease to produce hormone-refractory cancer cells renders this an ineffective long-term treatment. Many studies have suggested that the androgen receptor (AR) plays a critical role in the development of these hormone-insensitive cancer cells. Dr. Ingo Mellinghoff of the University of California, Los Angeles, has conducted studies that point to an additional level of complexity regarding how the AR does its job. Dr. Mellinghoff's research, supported by a Fiscal Year 2003 Physician Research Training Award, has demonstrated that the activity of the AR is regulated by yet another receptor family, known as the Epidermal Growth Factor Receptor family, or ErbB receptors. This finding strongly supports the idea that other therapies targeting Her2/ErbB, such as the breast cancer drug Herceptin, may prove efficacious against prostate cancer progression.

Publications:

Mellinghoff, I.K., Vivanco, I., Kwon, A., Tran, C., Wongvipat, J., and Sawyers, C.L. 2004. HER2/neu kinase-dependent modulation of androgen receptor function through effects on DNA binding and stability. Cancer Cell 6:517-527.

Xin L, Teitell M.A, Lawson D.A., Kwon A., Mellinghoff I.K., and Witte O.N. 2006. Progression of prostate cancer by synergy of Akt with genotropic and nongenotropic actions of the androgen receptor. Proc Natl Acad Sci U S A 103, 7789-7794.

Link:

Abstract: Regulation of Androgen Receptor Function by ErbB Receptor Tyrosine Kinases

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Delivering a One-Two Punch to Prostate Cancer
Posted July 24, 2007
Jacqueline Moreno, Ph.D., Stanford University School of Medicine, Stanford, California

Recent studies suggest that combination therapy for cancer treatment offers enhanced efficacy at lower doses while minimizing toxicity-related side effects. Dr. Jacqueline Moreno and colleagues at Stanford University School of Medicine have demonstrated the promise of this approach for prostate cancer treatment. With the support of a Fiscal Year 2004 Prostate Cancer Research Program Postdoctoral Traineeship Award, Dr. Moreno examined the combined effects of two agents on prostate cancer proliferation. The first agent investigated was 1,25-dihydroxyvitamin D3 (calcitriol), the active form of vitamin D. Calcitriol has well-established antiproliferative and prodifferentiation effects in human prostate cancer cells. Dr. Moreno showed that the effects of calcitriol on prostate cancer cells are mediated, at least in part, by inhibiting the synthesis and growth-stimulatory activity of prostaglandins (PGs), signaling molecules that are implicated in prostate cancer initiation and progression. Specifically, calcitriol reduced expression of cyclooxygenase (COX)-2, the inducible isoform of COX that initiates PG synthesis. Conversely, calcitriol enhanced expression of NAD+-dependent 15-hydroxyprostaglandin dehydrogenase, a putative tumor suppressor that converts PGs into inactive metabolites. Further, calcitriol downregulated expression of the PG receptors EP2 and FP. These results suggested that coupling calcitriol with a second drug to inhibit COX-2 activity could provide a "one-two punch" that would substantially impair prostate cancer cell proliferation. Dr. Moreno assessed the antiproliferative effects of calcitriol in conjunction with various COX-1 and -2 inhibitors known as non-steroidal anti-inflammatory drugs (NSAIDs), many of which are under active investigation for cancer prevention and treatment. The combination of calcitriol with ibuprofen, naproxen, or other NSAIDs synergistically inhibited prostate cancer cell growth at approximately 2- to 10-fold lower concentrations than the individual drugs. Based on these results, Dr. Sandy Srinivas of Stanford University, in collaboration with Dr. David Feldman (Dr. Moreno's mentor) and Dr. Natalia Colocci, have initiated a Phase II clinical trial (not funded by the Department of Defense) to examine the efficacy of calcitriol and naproxen in men with advanced, relapsing prostate cancer. Such combined treatment strategies may offer new hope to prostate cancer patients for whom other therapies have not been successful.

Publications:

Moreno J, Krishnan AV, Swami S, Nonn L, Peehl DM, and Feldman D. 2005. Regulation of prostaglandin metabolism by calcitriol attenuates growth stimulation in prostate cancer cells. Cancer Research 65:7917-7925.

Moreno J, Krishnan AV, and Feldman D. 2005. Molecular mechanisms mediating the anti-proliferative effects of vitamin D in prostate cancer. The Journal of Steroid Biochemistry and Molecular Biology 97:31-36.

Moreno J, Krishnan AV, Peehl DM, and Feldman D. 2006. Mechanisms of vitamin D-mediated growth inhibition in prostate cancer cells: Inhibition of the prostaglandin pathway. Anticancer Research 26(4A):2525-2530.

Krishnan AV, Moreno J, Nonn L, Malloy P, Swami S, Peng L, Peehl DM, and Feldman D. 2007. Novel pathways that contribute to the anti-proliferative and chemopreventive activities of calcitriol in prostate cancer. The Journal of Steroid Biochemistry and Molecular Biology 103(3-5):694-702.

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The Power of Applied Knowledge: Using Proteomics and Molecular Biology to Develop the Next Generation of Prostate Cancer Biomarkers
Posted July 5, 2007
Youqiang Ke, Ph.D., University of Liverpool, Liverpool, United Kingdom

Accurate, early detection and diagnosis of prostate cancer remains elusive. The familiar prostate-specific antigen (PSA) test represents the first generation of biochemically-based tools for prostate cancer detection. While this test has undoubtedly revolutionized the detection and diagnosis of prostate cancer, the PSA test has proven non-specific for prostate cancer. Elevated PSA levels are observed in conditions such as benign prostatic hyperplasia (BPH) and prostatitis. Additionally, the inability of the test to distinguish among different types of early-stage tumors limits its usefulness for assessing tumor aggressiveness and guiding treatment decisions. Advances in the understanding of the molecular changes that drive prostate cancer, combined with state-of-the-art methods proteomics and molecular biology, offer promise for the identification of the next generation of biomarkers for prostate cancer detection and diagnosis. Dr. Youqiang Ke of the University of Liverpool, with funding from an FY04 PCRP Exploration-Hypothesis Award, has identified two new candidate biomarkers

Using microquantity differential display, Dr. Ke discovered that the ribosomal protein L19 (RPL19) gene was overexpressed in malignant prostate cells and tissues. Further studies revealed a correlation between RPL19 protein levels and Gleason score, a common system for grading prostate cancer pathological features and aggressiveness. Staining intensity was significantly stronger in highly malignant tissue than that in moderately malignant tissues. Also there was a unique staining pattern for conditions such as BPH. Finally, RPL19 was a powerful predictor of survival. Prostate cancer patients whose tumors stained weakly for RPL19 survived longer than patients with strongly staining tumors.

Dr. Ke also studied Osteopontin (OPN), a well-established tumor promoter. Using several prostate cancer cell lines, he found that the level of OPN in the malignant LNCaP cell line was higher than in benign PNT-2 cells. Subsequently, a total of 116 tissue samples were analyzed for OPN expression. No significant differences in OPN staining were observed between normal and BPH samples. Strikingly, the level of OPN staining in carcinoma tissues was significantly stronger than in either normal or BPH tissues. These results suggest that elevated OPN expression is closely associated with increased prostate cancer aggressiveness and that OPN is capable of distinguishing between BPH and prostate cancer. Finally, the degree of OPN staining inversely correlated with patient survival, which strongly bolsters the potential for OPN as a future biomarker for prostate cancer.

Dr. Ke's work has produced two promising markers capable of distinguishing advanced, aggressive forms of prostate cancer from benign disease. This work could augment the detection, diagnosis and treatment of prostate cancer

Selected Publications:

Shiva S. Forootan, Christopher S. Foster, Vijay R. Aachi, Janet Adamson, Paul H. Smith, Ke Lin and Youqiang Ke "Prognostic significance of osteopontin expression in human prostate cancer" Int. J. Cancer: 118, 2255-2261 (2006)

Alix Bee, Youqianq Ke, Shiva Forootan, Ke Lin, Carol Beesley, Sharon E. Forrest and Christopher S. Foster "Ribosomal Protein L19 Is a Prognostic Marker for Human Prostate Cancer" Clin Cancer Res: 12(7) 2061-2065 (2006)

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Multiple Pathways Converge on BAD Protein to Aid Survival of Prostate Cancer Cells
Posted June 20, 2007
George Kulik, D.V.M., Ph.D., Wake Forest University School of Medicine, Winston-Salem, North Carolina

Prostate cancer cells are protected from cell death by different signaling pathways involving vasoactive intestinal peptide (VIP), epidermal growth factor (EGF), or phosphatidylinositol 3-kinase (PI3K). However, the intricacies of the pathways they use and the molecules they target are unknown. With studies funded by a Department of Defense Prostate Cancer Research Program Fiscal Year 2001 New Investigator Award, Dr. George Kulik of the Wake Forest University School of Medicine and colleagues found that all three signaling pathways work by inactivating a proapoptotic protein known as BAD, a member of the Bcl-2 family. PI3K, EGF, and VIP exert their anti-apoptotic effect by phosphorylating BAD. BAD phosphorylation is necessary for cancer cell survival mediated by these three agents. They also showed that the three agents employ different routes to phosphorylate BAD and achieve cancer cell survival. BAD phosphorylation induced by EGF is mediated by Ras/MEK module or Rac/PAK1 signaling, VIP effect is mediated by protein kinase A, and PI3K employs AktI as the principal kinase.

The investigators also are the first to establish the connection between stress and cancer. They found that the stress hormone, epinephrine, also protects prostate cancer cells from apoptosis through BAD phosphorylation by protein kinase A. Moreover, epinephrine exerts its effect at concentrations found after acute and chronic psychological stress. Thus, PI3K, EGF, VIP, and epinephrine protect prostate cancer cells from apoptosis by independent signaling pathways, all of which converge on BAD protein.

Since several agents and pathways aid prostate cancer cell survival through BAD phosphorylation, direct targeting of BAD is a promising therapeutic goal to regulate the apoptosis of prostate cells.

Publications:

Sastry KS, Karpova Y, Prokopovich S, Smith AJ, Essau B, Gersappe A, Carson JP, Weber MJ, Register TC, Chen YQ, Penn RB, and Kulik G. 2007. Epinephrine protects cancer cells from apoptosis via activation of cAMP-dependent protein kinase and BAD phosphorylation. Journal of Biological Chemistry 282(19):14094-14100.

Sastry KS, Smith AJ, Karpova Y, Datta SR, and Kulik G. 2006. Diverse antiapoptotic signaling pathways activated by vasoactive intestinal polypeptide, epidermal growth factor, and phosphatidylinositol 3-kinase in prostate cancer cells converge on BAD. Journal of Biological Chemistry 281(30):20891-20901.

Sastry KS, Karpova Y, and Kulik G. 2006. Epidermal growth factor protects prostate cancer cells from apoptosis by inducing BAD phosphorylation via redundant signaling pathways. Journal of Biological Chemistry 281(37):27367-27377.

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Maspin Suppresses Tumor Progression by Direct Interaction with Glutathione-S-Transferase
Posted May 14, 2007
Shuping Yin, Ph.D., Wayne State University School of Medicine, Detroit, Michigan

Maspin, a serine protease inhibitor, suppresses tumor progression in several cancer models, including prostate cancer. Maspin exerts its effects through intracellular and extracellular mechanisms. However, the specific mechanisms of action of maspin and its molecular targets are largely unknown. Considering the therapeutic potential of maspin, Dr. Shuping Yin of Wayne State University undertook a study, funded by a Department of Defense Prostate Cancer Research Program Fiscal Year 2002 Postdoctoral Traineeship Award, to characterize the intracellular and extracellular molecular mechanisms of action of maspin. Using a yeast two-hybrid system, Dr. Yin and colleagues showed that maspin interacts specifically with the intracellular protein glutathione-S-transferase (GST). A maspin variant that has a point mutation of Arg340 to Ala showed a significantly reduced affinity for GST. The authors also found that maspin expression in prostate cancer cells correlated with decreased basal levels of reactive oxygen species (ROS). Furthermore, the maspin effect on ROS generation was completely abolished by a synthetic GST inhibitor, ethacrynic acid. Thus, maspin mediates a cellular response to oxidative stress through its interaction with GST. Maspin inhibition of ROS may also lead to downstream effects on angiogenesis, since vascular endothelial growth factor A (VEGF-A) expression induced by oxidative stress is significantly reduced in prostate cancer cells expressing maspin. The data suggest an important role of intracellular maspin in the cellular response to oxidative stress and in tumor vascularization. Dr. Yin's research team also explored the extracellular mechanisms of maspin and found that secreted maspin inhibited the calcium reduction-induced detachment of prostate cancer cells. This effect was associated with increased and sustained levels of mature focal adhesion contacts (FACs). Furthermore, this effect on FACs was dependent on the interaction between urokinase-type plasminogen activator (uPA) and uPA receptor (uPAR), which was facilitated by extracellular maspin through its reactive site loop. These results provide the first evidence that extracellular maspin strengthens mature FACs and retards cell detachment through cooperation with the uPA-uPAR complex. Taken together, the data from Dr. Yin's team identify multiple actions of maspin in angiogenesis and tumor cell detachment, both of which are critical points in cancer progression, and support a rationale for developing new maspin-based therapeutics.

Publications

Yin S, Lockett J, Meng Y, et al. 2006. Maspin retards cell detachment via a novel interaction with the urokinase-type plasminogen activator/urokinase-type plasminogen activator receptor system. Cancer Research 66:(8):4173-4181.

Yin S, Li X, Meng Y, et al. 2005. Tumor-suppressive maspin regulates cell response to oxidative stress by direct interaction with glutathione S-transferase. Journal of Biological Chemistry280:34985-34996.

Link for Dr. Gelmann and Dr. Huang:

Abstract: The Molecular Basis for Maspin-Sensitized Prostate Epithelial Cell Apoptosis

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