Sowmyalakshmi Srinivasan

Par-4-Dependent Apoptosis by the Dietary Compound Withaferin A in Prostate Cancer Cells

Deletion or mutation of the androgen receptor (AR) renders prostate tumors refractory to apoptosis by androgen ablation, the mainstay of prostate cancer therapy. To identify novel therapeutics that can induce apoptosis regardless of the AR status of prostate cancer cells, we screened dietary herbal compounds using a reporter assay for the prostate apoptosis response-4 (Par-4) gene, which induces p53- and PTEN-independent and cancer-selective apoptosis. One of the compounds, withaferin A (WA), a major constituent of the dietary compound Withania somnifera, induced Par-4-dependent apoptosis in androgen-refractory prostate cancer cells and regression of PC-3 xenografts in nude mice. Interestingly, restoration of wild-type AR in PC-3 (AR negative) cells abrogated both Par-4 induction and apoptosis by WA. Individually, WA and anti-androgens induced neither Par-4 nor apoptosis in androgen-responsive prostate cancer cells, yet in combination, WA and anti-androgen synergistically induced Par-4 and apoptosis in androgen-responsive prostate cancer cells. Thus, when judiciously combined with anti-androgens, WA inhibits survival of both androgen-responsive and androgen-refractory prostate cancer cells by a Par-4-dependent mechanism. As Par-4 up-regulation induces apoptosis in most tumor cells, our findings can be extended to high-throughput screens to identify synergistic combinations for both therapy-sensitive and therapy-resistant cancers.

Notch-1 inhibition by Withaferin-A: a therapeutic target against colon carcinogenesis.

Notch signaling plays a crucial role in the development of colon cancer; targeting the Notch pathway may sensitize colon cancers to various adjuvant agents. The focus of our current study is to identify natural compounds that target Notch signaling and that might be beneficial for the prevention and treatment of colon cancer. Withaferin-A (WA) is a bioactive compound derived from Withania somnifera, which inhibits Notch-1 signaling and downregulates prosurvival pathways, such as Akt/NF-κB/Bcl-2, in three colon cancer cell lines (HCT-116, SW-480, and SW-620). In addition, WA downregulated the expression of mammalian target of rapamycin signaling components, pS6K and p4E-BP1, and activated c-Jun-NH2-kinase–mediated apoptosis in colon cancer cells. We also established the molecular link between Notch/Akt/mammalian target of rapamycin signaling by complementary approaches (i.e., overexpression of Notch-1 or inhibition of Notch-1 by small interfering RNA). Our results suggest that WA inhibits Notch-mediated prosurvival signaling, which facilitates c-Jun-NH2-kinase–mediated apoptosis in colon cancer cell lines. These results underscore the anticancer activity of WA, which exhibits potential for further development for targeted chemotherapy and/or chemoprevention strategies in the context of colon cancer. Mol Cancer Ther; 9(1); 202–210

Diosgenin targets Akt-mediated prosurvival signaling in human breast cancer cells.

In recent years, Akt signaling has gained recognition for its functional role in more aggressive, therapy‐resistant malignancies. As it is frequently constitutively active in cancer cells, several drugs are being investigated for their ability to inhibit Akt signaling. The purpose of this study is to determine effect of diosgenin (fenugreek), a dietary compound on Akt signaling and its downstream targets on estrogen receptor positive (ER+) and estrogen receptor negative (ER−) breast cancer (BCa) cells. Diosgenin inhibits pAkt expression and Akt kinase activity without affecting PI3 kinase levels, resulting in the inhibition of its downstream targets, NF‐κB, Bcl‐2, survivin and XIAP. The Raf/MEK/ERK pathway, another functional downstream target of Akt, was inhibited by diosgenin in ER+ but not in ER− BCa cells. Additionally, we found that diosgenin caused G1 cell cycle arrest by downregulating cyclin D1, cdk‐2 and cdk‐4 expression in both ER+ and ER− BCa cells resulting in the inhibition of cell proliferation and induction of apoptosis. Interestingly, no significant toxicity was seen in the normal breast epithelial cells (MCF‐10A) following treatment with diosgenin. Additionally, in vivo tumor studies indicate diosgenin significantly inhibits tumor growth in both MCF‐7 and MDA‐231 xenografts in nude mice. Thus, these results suggest that diosgenin might prove to be a potential chemotherapeutic agent for the treatment of BCa.

Landomycins P-W, cytotoxic angucyclines from Streptomyces cyanogenus S-136.

Streptomyces cyanogenus S-136 is the producer of previously reported landomycins A-D. An analysis of minor products of the strain led to isolation and structure elucidation of eight new congeners, named landomycins P-W (5, 6, 3, 17, 9, 10, 15, 7), along with 10 other known angucyclin(on)es. The structures of the new compounds were established from their NMR and mass spectrometry data. The activity of these angucyclin(on)es was determined using MCF-7 (estrogen responsive) and MDA-231 (estrogen refractory) breast cancer cell lines. Cell viability assays showed that anhydrolandomycinone (2), landomycinone (11), and landomycin A (16) showed the best combined activities in both MCF-7 and MDA-231 assays, with 2 being the most potent in the former and 11 and 16 in the latter. These data reveal that some of the aglycones are equipotent to the principle product 16, which contains the longest saccharide chain. Specifically, anhydrolandomycinone (2) was the most active against MCF-7 cells (IC(50) = 1.8 μM). Compounds with shorter saccharidal moieties were less potent against MCF-7. The fact that the most active landomycins have either long penta- or hexasaccharide chains or no sugars at all suggests that the large compounds may act by a different mode of action than their small sugar-free congeners. The results presented here provide more insights into the structure-activity relationship of landomycins.

Psoralidin, an Herbal Molecule, Inhibits Phosphatidylinositol 3-Kinase–Mediated Akt Signaling in Androgen-Independent Prostate Cancer Cells

The protein kinase Akt plays an important role in cell proliferation and survival in many cancers, including prostate cancer. Due to its kinase activity, it serves as a molecular conduit for inhibiting apoptosis and promoting angiogenesis in most cell types. In most of the prostate tumors, Akt signaling is constitutively activated due to the deletion or mutation of the tumor suppressor PTEN, which negatively regulates phosphatidylinositol 3-kinase through lipid phosphatase activity. Recently, we identified a natural compound, psoralidin, which inhibits Akt phosphorylation, and its consequent activation in androgen-independent prostate cancer (AIPC) cells. Furthermore, ectopic expression of Akt renders AIPC cells resistant to chemotherapy; however, psoralidin overcomes Akt-mediated resistance and induces apoptosis in AIPC cells. While dissecting the molecular events, both upstream and downstream of Akt, we found that psoralidin inhibits phosphatidylinositol 3-kinase activation and transcriptionally represses the activation of nuclear factor-κB and its target genes (Bcl-2, Survivin, and Bcl-xL, etc.), which results in the inhibition of cell viability and induction of apoptosis in PC-3 and DU-145 cells. Interestingly, psoralidin selectively targets cancer cells without causing any toxicity to normal prostate epithelial cells. In vivo xenograft assays substantiate these in vitro findings and show that psoralidin inhibits prostate tumor growth in nude mice. Our findings are of therapeutic significance in the management of prostate cancer patients with advanced or metastatic disease, as they provide new directions for the development of a phytochemical-based platform for prevention and treatment strategies for AIPC.

Inhibiting TNF-mediated signaling: a novel therapeutic paradigm for androgen independent prostate cancer.

The tumor necrosis factor (TNF) receptor super family comprises of members that induce two distinct signaling cascades, leading to either cell survival or apoptosis. However, in prostate cancer (PCa), TNF-mediated prosurvival signaling is the predominant pathway that leads to cell survival and resistance to therapy. Although inhibition of TNF signaling by pharmacological agents or monoclonal antibodies has gained importance in the field of cancer therapy, toxicity to normal cells has impaired their extensive use for cancer treatment. We previously identified a natural, nontoxic compound psoralidin that inhibited viability and induced apoptosis in androgen independent prostate cancer (AIPC) cells. Thus, the goal of our study is to investigate whether psoralidin inhibits TNF-mediated prosurvival signaling in AIPC cells. Our results suggest that psoralidin inhibits constitutive and TNF-induced expression of TNF-α and its downstream prosurvival signaling molecules such as NF-κB and Bcl-2 in AIPC cells. On the other hand, psoralidin simultaneously induces the death receptor (DR)-mediated apoptotic signaling eventually causing the activation of caspase cascade and resultant induction of apoptosis. Oral administration of psoralidin inhibits expression of TNF-α and NF-κB/p65 in tumor sections, resulting in tumor regression in PC-3 xenografts. Our results suggest that psoralidin inhibits TNF-mediated survival signaling in AIPC and thus is a potent therapeutic agent for prostate cancer.

Activating Stress-Activated Protein Kinase–Mediated Cell Death and Inhibiting Epidermal Growth Factor Receptor Signaling: A Promising Therapeutic Strategy for Prostate Cancer

Epidermal growth factor receptor (EGFR) activation is an important event that regulates mitogenic signaling, such as the Raf, mitogen-activated protein kinase (MAPK), and extracellular signal–regulated kinase 1/2 cascades. EGFR activation has been implicated in the transition of prostate cancer from androgen dependence to independence. Therefore, inhibition of EGFR may effectively suppress prostate cancer growth and progression. The goal of this study was to determine whether the natural compound psoralidin alters EGFR-mediated signaling resulting in the inhibition of prostate cancer growth. Results suggest that inhibition of EGFR alone (by serum deprivation) fails to induce stress-mediated protein kinases (SAPK), namely, Jun NH2-terminal kinase/c-Jun signaling, in androgen-independent prostate cancer (AIPC) cells. Treatment with psoralidin, however, inhibited both constitutive and EGF-induced EGFR activation and simultaneously triggered SAPK signaling, resulting in the induction of apoptosis in AIPC cells. In addition, psoralidin downregulated EGFR-regulated MAPK signaling and inhibited cell proliferation in AIPC cells. Oral administration of psoralidin effectively suppressed PC-3 xenograft tumors in nude mice. Compared with control tumors, inhibition of pEGFR expression and an increase in the phosphorylation, activation, and nuclear translocation of c-Jun were observed in psoralidin-treated tumor sections. Our studies suggest that psoralidin may be a potent therapeutic agent that modulates EGFR-mediated key epigenetic events in AIPC. Mol Cancer Ther; 9(9); 2488–96. ©2010 AACR.

Identification of urushiols as the major active principle of the Siddha herbal medicine Semecarpus Lehyam: Anti-tumor agents for the treatment of breast cancer

Breast cancer (BCa) is the most commonly occurring cancer in women, comprising almost one third of all malignancies. Previously we reported that the n-hexane fraction (hSL) of the Siddha herbal medicine, Semecarpus Lehyam, relatively sensitized estrogen receptor-negative (ER−) BCa when compared to estrogen receptor-positive (ER+) BCa cells. In this study we used a bioassay-guided fractionation approach leading to a simplified fraction of hSL that effectively sensitized both ER+ (MCF-7) and ER− (MDA-231) BCa cells. Further bioassay-guided isolation led to the purification of three potent anti-cancer components from hSL which significantly induced apoptosis in both the BCa cell lines. Their structures were identified through NMR and mass spectroscopic analysis as (7;Z,10;Z)-3-pentadeca-7,10-dienyl-benzene-1,2-diol (1), (8;Z)-3-pentadec-10-enyl-benzene-1,2-diol (2) and 3-pentadecyl-benzene-1,2-diol (3). Compounds (1) and (2) turned out to be more active than (3). The overall results of this study suggest that these major components of hSL may be solely responsible for the anti-tumor effect of SL.

Abstract 4043: Targeting microRNA for the prevention and treatment of prostate cancer

MicroRNAs (miRs) are small 20-24 nucleotide non-coding RNA molecules that are involved in post-transcriptional regulation of gene expression in multicellular organisms by affecting both stability and translation of mRNAs. The expression and activity of miRs vary in specific cell types and disease states. Several groups have investigated and determined the strong association between specific expression patterns of miRs and various stages of prostate cancer. Recent findings have highlighted the strong correlation between overexpression of miRs (21, 210, 125b, 221 and 222) and cancer development and progression. In this study we determined whether modulation of miR expression using a natural compound suppresses PCa growth. Previously we published that psoralidin inhibits prostate tumor growth and progression in both in vitro and in vivo models. While studying the expression patterns of a panel of miRs in control and psoralidin treated prostate cancer cells we found that psoralidin significantly inhibits expression of two specific miRNAs, miR-21 and miR-210 in turn resulting in the inhibition of EGFR-, MAPK- and PI3K-mediated prosurvival signaling and induction of JNK-mediated apoptosis in prostate cancer cells. Currently, we are overexpressing and inhibiting these two miRs to determine whether it modulates psoralidin-mediated effects on prostate cancer cells. The outcome of our study may provide encouraging data that may enable us to exploit miRs as therapeutic targets for the treatment of various malignancies. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. 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 4043.