We demonstrate, for the first time, that the transcription factor NF-jB is constitutively activated during human cervical cancer progression. Immunohistochemical analysis was done using 106 paraffin-embedded cervical tissue specimens of different histological grades. In normal cervical tissue and low-grade squamous intraepithelial lesions, p50, RelA and IjB-a were mainly localized in the cytosol, whereas in high-grade lesions and squamous cell carcinomas, p50-RelA heterodimers translocated into the nucleus with a concurrent decrease in IjB-a protein. By Western blot analysis, p50 and RelA were detectable mainly in the cytosolic and nuclear extracts in normal and cancer tissues, respectively, and cytosolic IjB-a expression was detectable in normal but not in cancer cervical tissues. NF-jB DNA-binding activity increased during cervical cancer progression and the binding complex was mainly composed of the p50-RelA heterodimers as revealed by electrophoretic mobility shift assays. Semiquantitative RT-PCR analysis, however, showed increased levels of IjB-a mRNA in cancer samples presumably because of feedback regulation as a result of enhanced NF-jB DNA-binding activity and a consequent functional activation of NF-jB. Further immunohistochemical analysis with an antibody to phospho IjB-a revealed that phosphorylation occurs mainly in squamous intraepithelial lesions, suggesting that the IjB-a gets phosphorylated initially and degraded as the tumor progressed.

This review provides an overview of the clinical relevance of chemosensitization, giving special reference to the phenolic phytochemicals, curcumin, genistein, epigallocatechin gallate, quercetin, emodin, and resveratrol, which are potential candidates due to their ability to regulate multiple survival pathways without inducing toxicity. We also give a brief summary of all the clinical trials related to the important phytochemicals that emerge as chemosensitizers. The mode of action of these phytochemicals in regulating the key players of the death receptor pathway and multidrug resistance proteins is also abridged. Rigorous efforts in identifying novel chemosensitizers and unraveling their molecular mechanism have resulted in some of the promising candidates such as curcumin, genistein, and polyphenon E, which have gone into clinical trials. Even though considerable research has been conducted in identifying the salient molecular players either contributing to drug efflux or inhibiting DNA repair and apoptosis, both of which ultimately lead to the development of chemoresistance, the interdependence of the molecular pathways leading to chemoresistance is still the impeding factor in the success of chemotherapy. Even though clinical trials are going on to evaluate the chemosensitizing efficacy of phytochemicals such as curcumin, genistein, and polyphenon E, recent results indicate that more intense study is required to confirm their clinical efficacy. Current reports also warrant intense investigation about the use of more phytochemicals such as quercetin, emodin, and resveratrol as chemosensitizers, as all of them have been shown to modulate one or more of the key regulators of chemoresistance.

No abstract

No abstract

Human epithelial tumor progression and metastasis involve cellular invasion, dissemination in the vasculature, and regrowth at metastatic sites. Notch signaling has been implicated in metastatic progression but its roles have yet to be fully understood. Here we report the important role of Notch signaling in maintaining cells expressing the carcinoembryonic antigen cell adhesion molecule CEACAM (CD66), a known mediator of metastasis. CD66 and Notch1 were studied in clinical specimens and explants of human cervical cancer, including specimens grown in a pathophysiologically relevant murine model. Gene expression profiling of CD66 þ cells from primary tumors showed enhanced features of Notch signaling, metastasis, and stemness.Significant differences were also seen in invasion, colony formation, and tumor forming efficiency between CD66 þ and CD66 À cancer cells. Notably, CD66 þ cells showed a marked sensitivity to a Notch small molecule inhibitor. In support of studies in established cell lines, we documented the emergence of a tumorigenic CD66 þ cell subset within a metastatic lesion-derived cervical-cancer cell line. Similar to primary cancers, CD66 expression in the cell line was blocked by chemical and genetic inhibitors of ligand-dependent nuclear Notch signaling. Collectively, our work on the oncogenic properties of CD66 þ cells in epithelial cancers provides insights into the nature of tumor progression and offers a mechanistic rationale to inhibit the Notch signaling pathway as a generalized therapeutic strategy to treat metastatic cancers. Cancer Res; 71(14); 4888-97. Ó2011 AACR.

No abstract

Curcumin (diferuloyl methane), the yellow pigment in turmeric (Curcuma longa), possesses anti-inflammatory and antioxidant activities (1, 2). It is also a potent chemopreventive agent inhibiting tumor promotion against skin, oral, intestinal, and colon carcinogenesis (3-5). Many chemopreventive agents induce programmed cell death or apoptosis, a potent mechanism by which they eliminate preneoplastic or cancer cells. Curcumin induces several characteristics of apoptosis such as cell shrinkage, chromatin condensation, and DNA fragmentation in cells from colon, kidney, blood, and liver of human origin and NIH3T3, erbB2-transformed NIH3T3, and fibrosarcoma cells of murine origin (6, 7). However, curcumin fails to induce apoptosis in some fibroblastic and leukemic cell lines, and inhibition of cell proliferation is not always associated with apoptosis (8 -10). Although the molecular mechanisms of antiproliferative and apoptotic effects of curcumin remain elusive, the resistance of various cell types to apoptosis following treatment with curcumin may be attributed to the abnormal expression of specific proteins that regulate apoptosis. Many cancer cells protect themselves against therapy (tumor necrosis factor, ionizing radiation, and chemotherapeutic compounds) by activating NF-B 1 /Rel, a family of dimeric transcription factors, that helps in cell survival through unknown anti-apoptotic mechanisms (11)(12)(13)(14). Interestingly, constitutive expression of NF-B has been reported in breast and pancreatic cancer cells and tumors (but not in their normal counterparts), contributing to their apoptosis resistance/survival and tumor progression (15, 16). The typically active NF-B/Rel is a heterodimer composed of a 50-kDa (p50) and a 65-kDa (p65 or RelA) subunit, although other homo/heterodimers of the mammalian Rel family (c-Rel, p52, and RelB) also exist. Knockout mice lacking the RelA subunit of NF-B die before birth and show massive degeneration of liver cells caused by apoptosis. NF-B proteins are sequestered in cytoplasm as inactive forms by associating with inhibitory proteins known as IBs. IB-␣ represents a prototype of several IB proteins and IB kinases, which phosphorylate them on specific serine residues upon getting a signal, have been cloned and characterized recently (17). Phosphorylation of IB-␣ leads to its ubiquitination and subsequent degradation eventually releasing the heterodimers of NF-B to translocate to the nucleus (18).Curcumin is known to inhibit the activation of NF-B induced by tumor necrosis factor-␣, phorbol esters, and hydrogen peroxide (19). Curcumin inhibits cytokine-mediated NF-B activation by blocking a signal leading to IB kinase activity (20). Presumably, curcumin-induced apoptosis is regulated, in part, by the extent of NF-B inactivation by curcumin and the level of expression of NF-B in cells. To understand the role of NF-B in curcumin-induced apoptosis, we stably transfected relA gene into L-929 cells (mouse fibrosarcoma), and upon treatment with curcumin, the relA-transfected cells ...

A subpopulation within cancer, known as cancer stem cells (CSCs), regulates tumor initiation, chemoresistance, and metastasis. At a closer look, CSCs show functional heterogeneity and hierarchical organization. The present review is an attempt to assign marker profiles to define the functional heterogeneity and hierarchical organization of CSCs, based on a series of single-cell analyses. The evidences show that analogous to stem cell hierarchy, self-renewing Quiescent CSCs give rise to the Progenitor CSCs with limited proliferative capacity, and later to a Progenitor-like CSCs, which differentiates to Proliferating non-CSCs. Functionally, the CSCs can be tumor-initiating cells (TICs), drug-resistant CSCs, or metastasis initiating cells (MICs). Although there are certain marker profiles used to identify CSCs of different cancers, molecules like CD44, CD133, ALDH1A1, ABCG2, and pluripotency markers [Octamer binding transcriptional factor 4 (OCT4), SOX2, and NANOG] are used to mark CSCs of a wide range of cancers, ranging from hematological malignancies to solid tumors. Our analysis of the recent reports showed that a combination of these markers can demarcate the heterogeneous CSCs in solid tumors. Reporter constructs are widely used for easy identification and quantification of marker molecules. In this review, we discuss the suitability of reporters for the widely used CSC markers that can define the heterogeneous CSCs. Since the CSC-specific functions of CD44 and CD133 are regulated at the post-translational level, we do not recommend the reporters for these molecules for the detection of CSCs. A promoter-based reporter for ABCG2 may also be not relevant in CSCs, as the expression of the molecule in cancer is mainly regulated by promoter demethylation. In this context, a dual reporter consisting of one of the pluripotency markers and ALDH1A1 will be useful in marking the heterogeneous CSCs. This system can be easily adapted to high-throughput platforms to screen drugs for eliminating CSCs.