Stages Of Cancer Development Research Articles

Hospicells promote upregulation of the ATP-binding cassette genes by insulin-like growth factor-I via the JAK2/STAT3 signaling pathway in an ovarian cancer cell line

Interaction between tumor cells and their microenvironment has a crucial role in the development, progression and drug resistance of cancer. Our objective was to confirm the role of Hospicells, which are stromal cells from the cancer microenvironment, in drug resistance and tumor cell growth. We demonstrated that soluble factors secreted by Hospicells activate several genes and upregulate the JAK/STAT signaling pathway in ovarian cancer cell lines. Hospicells express all insulin-like growth factor (IGF) family as detected by gene array, RT-PCR, protein array and immunocytochemistry. While focusing attention on the microenvironment, we considered the role of IGF-I in proliferation and survival of ovarian cancer cells. Indeed, IGF-I is a major regulator of different stages of cancer development. We studied the effect of exogenously added IGF-I on the regulation of ATP-binding cassette (ABC) genes (MDR1, MRP1, MRP2, MRP3, MRP5 and BCRP) in the ovarian cancer cell line OVCAR3 and validated the results obtained using the IGF-IR antagonist picropodophyllin. IGF-I regulates the expression of ABC genes in OVCAR3 cells via the PI3-kinase, MEK and JAK2/STAT3 signaling pathways. The OVCAR3 cell line when co-cultured with Hospicells showed a marked degree of drug resistance. The drug resistance observed could be amplified with exogenous IGF-I. Addition of IGF-IR inhibitor, however, reduced the degree of resistance in these exposed cells. Cells that were treated with anticancer drugs and then exposed to IGF-I showed an increase in drug resistance and, thereby, an increase in cell survival. This observation indicates that drug resistance of OVCAR3 cells increases when there is synergy between OVCAR3 cells and Hospicells and it is amplified when IGF-I was exogenously added. In conclusion, inhibition of IGF-IR and targeting of the JAK2/STAT3 signaling pathway can be a target for ovarian cancer therapy.

Abstract 5320: miR-211 expression inhibits MMP-9-induced phenotype and tumor growth in lung adenocarcinoma cancer stem cells in vitro and in vivo.

Abstract MicroRNAs are small non-coding RNAs that post-transcriptionally modulate gene expression by degrading or repressing the translation of target messenger RNAs (mRNA). MicroRNAs play fundamental roles in diverse biological and pathological processes, including cell proliferation, differentiation, apoptosis, and carcinogenesis. MicroRNAs have been revealed to be involved in various stages of cancer development, showing that abnormal miRNA expression plays a critical role in modulating expression of known oncogenes or tumor suppressor genes during cancer progression. In the present study, we observed that miR-211 expression was significantly lower in human non-small cell lung cancer (NSCLC) than their matched normal tissue samples, and we elucidated the tumor suppressor role of miR-211 using in vitro and in vivo models. We demonstrated that the expression of precursor of miR-211 (p-mir-211) inhibits radiation-induced invasive nature and tumor progression of NSCLC-initiating cells (NSCLC-ICs) in vitro and in vivo. Overexpression of miR-211 in NSCLC-ICs inhibited radiation-induced MMP-9 expression and invasive nature. Overexpression of miR-211 also inhibited epithelial to mesenchymal transition (EMT), which caused a decline in the mesenchymal markers vimentin, beta catenin and TGF-beta signaling molecules while increasing the epithelial marker, E-cadherin. Further, miR-211-treatment inhibited pre-established tumors in orthotopic mouse models. In addition, MMP-9 immunoreactivity decreased in tumor sections from mice that received miR-211 treatment as compared to tumor sections from mice that received control treatments. Finally, the apoptotic population was significantly increased in tumor sections from mice that received miR-211 treatment. Our data suggest miR-211 as a novel approach for the inhibition of radiation-induced invasion and tumor growth in NSCLC-ICs. Citation Format: Chandramu Chetty, Subramanyam Chittivelu, Jasti S. Rao. miR-211 expression inhibits MMP-9-induced phenotype and tumor growth in lung adenocarcinoma cancer stem cells in vitro and in vivo. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5320. doi:10.1158/1538-7445.AM2013-5320

Abstract 2158: Evaluation of the efficacy of dasatinib, a Src/Abl inhibitor, in a preclinical model of colorectal cancer.

Abstract Background: Dysregulation of c-Src has been shown to modulate many different tumor cell processes. Downstream targets of Src include focal adhesion kinase (FAK), which is important for migration and invasion, and Stat-3, a kinase that enhances survival of cells and angiogenesis. Dysregulation of Src has been shown to be important at various stages of cancer development. Dasatinib is a potent Src/Abl inhibitor and has been shown to have anti-proliferative and anti-invasive activity in many preclinical models. The objective was to determine the anti-proliferative and anti-tumor effects of dasatinib in our preclinical model of colorectal cancer (CRC). Experimental Design: We used colorectal cancer cell lines and patient-derived explants to investigate the efficacy of dasatinib. We treated 45 CRC cell lines with dasatinib for 72 hours and proliferation was assayed by a sulforhodamine B (SRB) assay; an IC50 < 0.1 μM was considered sensitive. We treated 10 patient-derived CRC explants with dasatinib (50 mg/kg/day, administered once-daily) for 28 days to determine in vivo efficacy; a tumor growth inhibition inhibition (TGI) ≥ 50% was considered sensitive. Results: We found that 17 out of 41 CRC cell lines reached an IC50 < 1 uM with dasatinib treatment. The cell lines: HCT116, LS174T, SW1417, WiDR, KM20 and SNU977 all reached an IC50 < 0.08 μM, which is considered to be an achievable dose in the clinic setting. Of 10 CRC explants, grown as xenografts, one showed sensitivity to dasatinib, with a tumor growth inhibiton of 55%. Conclusion: Dasatinib has significant anti-proliferative activity in a subset of CRC cell lines in vitro, but shows moderate efficacy in one out of ten explants treated. We plan on treating an additional 10 CRC explants (20 total) with dasatinib and developing biomarkers of sensitivity or resistance. Citation Format: Eun-Kee Song, Alicia Purkey, Kevin Quackenbush, Benjamin Cross, Stacey Bagby, John J. Arcaroli, Wells Messersmith. Evaluation of the efficacy of dasatinib, a Src/Abl inhibitor, in a preclinical model of colorectal cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2158. doi:10.1158/1538-7445.AM2013-2158

Abstract 5367: Comparison of global DNA methylation in uterine tissue from different species of monkeys and humans exposed to tamoxifen.

Abstract Tamoxifen (TAM) is a selective estrogen receptor modulator used worldwide for adjuvant therapy and chemoprevention of breast cancer. However, women receiving TAM therapy have an increased risk of endometrial and myometrial cancer, which may be due to genotoxicity and/or estrogen receptor-related mechanisms. It is now accepted that cancer can be both a genetic and an epigenetic disease, with these components participating at all stages of cancer development. DNA methylation at the 5-position of cytosine silences gene expression. Aberrant global DNA 5-methylcytosine (5-mC) levels may produce critical changes in the expression of genes that regulate cell growth and invasiveness. In order to study the effect of TAM exposure on uterine 5-mC levels, we examined uterine tissues taken from aging Erythrocebus patas (patas) and Macaca fascicularis (macaque) monkeys given oral TAM dosing for 3-4 months. In addition, we evaluated endometrial and myometrial samples from breast cancer survivors, who typically receive adjuvant TAM therapy for 5 years. The percentage of 5-mC in DNA was determined by an ELISA that employed a highly-specific 5-mC antibody. Of 5 female patas, 2 were unexposed and 3 were given oral dosing with 1.7 mg TAM/kg bw/day for 3 months. Of 12 female macaques, 6 were unexposed and 6 were given 1.3 mg TAM/kg bw/day for 4 months. Also, uterine tissue was taken at surgery or autopsy from normal-appearing areas and malignant-appearing areas, from women who received 20 mg TAM/day (n=9) and unexposed women (n=6). The patas monkeys given TAM had significantly increased levels of uterine 5-mC compared to the controls (2.9±0.13% vs. 1.9±0.26%, mean ± SE, respectively, p=0.03). However, the macaques showed no significant differences in uterine 5-mC between unexposed and TAM-exposed animals (0.93%±0.1 vs. 1.07%±0.05, mean ± SE, respectively). In human patients, TAM treatment did not alter the global 5-mC levels in either normal-appearing or malignant-appearing endometrium. However, global 5-mC levels in normal-appearing endometrium from the combined TAM-treated and untreated women (n=6) were significantly lower than those found in malignant-appearing uterine tissue from the combined TAM-treated and untreated (n=9) group (0.33%±0.01 vs. 0.70%±0.09, mean ± SE, respectively, p=0.006). Therefore, changes in uterine global DNA 5-mC are not altered consistently within primate species as a result of long-term TAM exposure, suggesting that 5-mC alterations are not major contributors to TAM-induced endometrial cancer. Continuing studies will evaluate 5-mC levels in the promoter regions of specific genes that are modulated in expression level during TAM therapy. Citation Format: Elena E. Hernandez Ramon, Nancy Si, Mark J. Cline, Charles E. Wood, Ruth Woodward, Miriam C. Poirier. Comparison of global DNA methylation in uterine tissue from different species of monkeys and humans exposed to tamoxifen. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5367. doi:10.1158/1538-7445.AM2013-5367

Importance of Serum SELDI-TOF-MS Analysis in the Diagnosis of Early Lung Cancer

Different methods of diagnosis have been found to be inefficient in terms of screening and early diagnosis of lung cancer. Cancer cells produce proteins whose serum levels may be elevated during the early stages of cancer development. Therefore, those proteins may be recognized as potential cancer markers. The aim of this study was to differentiate healthy individuals and lung cancer cases by analyzing their serum protein profiles and evaluate the efficacy of this method in the early diagnosis of lung cancer. 170 patients with lung cancer, 53 under high risk of lung cancer, and 47 healthy people were included in our study. Proteomic analysis of the samples was performed with the SELDI-TOF-MS approach. The most discriminatory peak of the high risk group was 8141. When tree classification analysis was performed between lung cancer and the healthy control group, 11547 was determined as the most discriminatory peak, with a sensitivity of 85.5%, a specificity of 89.4%, a positive predictive value (PPV) of 96.7% and a negative predictive value (NPV) of 62.7%. We determined three different protein peaks 11480, 11547 and 11679 were only present in the lung cancer group. The 8141 peak was found in the high-risk group, but not in the lung cancer and control groups. These peaks may prove to be markers of lung cancer which suggests that they may be used in the early diagnosis of lung cancer.

Two initiation sites of early detection of colon cancer revealed by localization of pERK1/2 in the nuclei or in aggregates at the perinuclear region of the tumor cells

We have used human specimens and antibodies to pERK1/2 to detect early development of colon cancer using indirect immunocytochemistry. Two distinct sites were stained; one at the tip of the colon crypts and the other in the stromal tissue associated with the colonic tissue. These foci represent early stages of colon cancer initiation sites as established by enhanced Kirsten Rat Sarcoma Virus (KRAS) and the lack of p53 staining. The enhanced KRAS coincides with the initiation of tumor growth revealed by pERK1/2, both in the tip of the colon crypts, as well as in the stromal initiation site of the colon tumors. Foci of pERK1/2 staining were also detected in 50% of stromal tissue and tips of colon crypts, which were classified as normal tissues, adjacent to the malignant tissue according to general morphology. However, in colon specimens, where no malignancy was observed, no accumulation of pERK1/2 was observed. The staining of pERK1/2 at the stromal foci of the apparently non-malignant tissue appeared as aggregates in the perinuclear region, while in the colon epithelium it appeared in the cell nuclei. In low-grade colon cancer that was still free of induced mutated p53, staining of pERK1/2 was prominent in the cell nuclei, both in the stroma tissue and the tip of the colon crypts. In the intermediate stage, that exhibited significant p53 staining, only a fraction of p53-free tumor cells was labeled with pERK1/2 antibody, while in high-grade tumors, all cells of tumors were labeled with antibodies to p53, but not with antibodies to pERK1/2. We suggest that the down regulation in pERK1/2 labeling is due to the mitogenic capacity of the tumor cells, which are shifted from being driven by nuclear pERK1/2 to mutated p53 expression. We also found that the cytoplasm of low grade tumors was positive for epiregulin, while this labeling decreased in high-grade tumors. We found that the tumors arising from the stroma demonstrated poor structural differentiation, while the tumors initiating from the epithelial cells of the colon demonstrated high structural differentiation. We conclude that pERK1/2 is a sensitive marker of early colon cancer, which disappears at later stages of cancer development. Moreover, pERK1/2 staining can distinguish between tumor cells originating from the tip of the colon crypts and those developing in the stroma, which is present in the close vicinity to colon epithelial tissue, and thus can assist in selecting the appropriate therapy.

Mesenchymal and stem-like cell properties targeted in suppression of chronically-induced breast cell carcinogenesis

Stem-like cells and the epithelial-to-mesenchymal transition (EMT) program are postulated to play important roles in various stages of cancer development, but their roles in breast cell carcinogenesis and intervention remain to be clarified. We investigated stem-like cell- and EMT-associated properties and markers in breast epithelial cells chronically exposed to low-dose 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and benzo[a]pyrene in the presence and absence of the preventive agents green tea catechins and grape seed extract. Our results indicate that stem-like cell- and EMT-associated properties and markers should be seriously considered as new cancer-associated indicators for detecting breast cell carcinogenesis and as endpoints for intervention of carcinogenesis.

Design, Synthesis, and Functional Evaluation of Leukocyte Function Associated Antigen-1 Antagonists in Early and Late Stages of Cancer Development

The integrin leukocyte function associated antigen 1 (LFA-1) binds the intercellular adhesion molecule 1 (ICAM-1) by its α(L)-chain inserted domain (I-domain). This interaction plays a key role in cancer and other diseases. We report the structure-based design, small-scale synthesis, and biological activity evaluation of a novel family of LFA-1 antagonists. The design led to the synthesis of a family of highly substituted homochiral pyrrolidines with antiproliferative and antimetastatic activity in a murine model of colon carcinoma, as well as potent antiadhesive properties in several cancer cell lines in the low micromolar range. NMR analysis of their binding to the isolated I-domain shows that they bind to the I-domain allosteric site (IDAS), the binding site of other allosteric LFA-1 inhibitors. These results provide evidence of the potential therapeutic value of a new set of LFA-1 inhibitors, whose further development is facilitated by a synthetic strategy that is versatile and fully stereocontrolled.

Is the Low-Resolution Mass Spectrometer Capable of Detecting Cancer at Early Stages?

It is known that cancer promotes changes in the metabolites found in biofluids and, therefore, the early detection of these metabolites allows for the best prognosis of the disease. Since ESI-MS fingerprinting has proved to be an excellent tool for ‘‘first pass’’ metabolome analysis of complex biological samples, we have presented, for the first time, the use of direct infusion electrospray ionization low-resolution mass spectrometry associated with chemometric analysis (ESI-DIMS-PCA) for the detection of early stage cancer in mice. Ehrlich tumors were induced in Balb-C and Swiss mice, and blood was collected 1, 2, 3, 5, 7 and 10 days after tumor induction. Data from the mass spectra were analyzed via principal component analysis (PCA). Sample classifications were obtained within 3 days of tumor induction, which reflects a precocious time for cancer detection. ESI-DIMS-PCA was also used to detect cancer in groups of Swiss mice inoculated with sarcoma 180. Sample classifications were obtained 3 days after tumor induction, which indicates ESI-DIMS-PCA can be used to detect distinct neoplasms. Loading plot analysis demonstrated that the ions differentiating between the experimental and control groups are lysophosphatidylcholines and phosphatidylcholines. Partial least-squares analysis demonstrated that ESI-DIMS-PCA is capable of determining unknown samples in different stages of cancer development. ESI-DIMS with chemometric analysis is a promising technique for diagnosing cancer in the early stages without the need for invasive procedures.

Role of inflammasomes and their regulators in prostate cancer initiation, progression and metastasis

Prostate cancer is one of the main cancers that affect men, especially older men. Though there has been considerable progress in understanding the progression of prostate cancer, the drivers of its development need to be studied more comprehensively. The emergence of resistant forms has also increased the clinical challenges involved in the treatment of prostate cancer. Recent evidence has suggested that inflammation might play an important role at various stages of cancer development. This review focuses on inflammasome research that is relevant to prostate cancer and indicates future avenues of study into its effective prevention and treatment through inflammasome regulation. With regard to prostate cancer, such research is still in its early stages. Further study is certainly necessary to gain a broader understanding of prostate cancer development and to create successful therapy solutions.

Flow-based pipeline for systematic modulation and analysis of 3D tumor microenvironments.

The cancer microenvironment, which incorporates interactions with stromal cells, extracellular matrix (ECM), and other tumor cells in a 3-dimensional (3D) context, has been implicated in every stage of cancer development, including growth of the primary tumor, metastatic spread, and response to treatment. Our understanding of the tumor microenvironment and our ability to develop new therapies would greatly benefit from tools that allow us to systematically probe microenvironmental cues within a 3D context. Here, we leveraged recent advances in microfluidic technology to develop a platform for high-throughput fabrication of tunable cellular microniches ("microtissues") that allow us to probe tumor cell response to a range of microenvironmental cues, including ECM, soluble factors, and stromal cells, all in 3D. We further combine this tunable microniche platform with rapid, flow-based population level analysis (n > 500), which permits analysis and sorting of microtissue populations both pre- and post-culture by a range of parameters, including proliferation and homotypic or heterotypic cell density. We used this platform to demonstrate differential responses of lung adenocarcinoma cells to a selection of ECM molecules and soluble factors. The cells exhibited enhanced or reduced proliferation when encapsulated in fibronectin- or collagen-1-containing microtissues, respectively, and they showed reduced proliferation in the presence of TGF-β, an effect that we did not observe in monolayer culture. We also measured tumor cell response to a panel of drug targets and found, in contrast to monolayer culture, specific sensitivity of tumor cells to TGFβR2 inhibitors, implying that TGF-β has an anti-proliferative affect that is unique to the 3D context and that this effect is mediated by TGFβR2. These findings highlight the importance of the microenvironmental context in therapeutic development and that the platform we present here allows the high-throughput study of tumor response to drugs as well as basic tumor biology in well-defined microenvironmental niches.

An Approach towards Automated Disease Diagnosis & Drug Design Using Hybrid Rough-Decision Tree from Microarray Dataset

Biological databases related to medical science, containing pathological, radiological and genetic information of patients is undergoing tremendous growth, beyond our analyzing capability. However such analysis can reveal new findings about the cause and subsequent treatment of any disease. Here the genetic information of Lung Adenocarcinoma, in the form of microarray dataset has been investigated which have five different stages. Rough Set Theory (RST) has been used in analysis with an aim to effectively extract biologically relevant information, as RST is a tool that works well in an environment, heavy with inconsistent and ambiguous data, or with missing data and provides efficient algorithms for finding hidden patterns in data. The investigation has been carried out on the publicly available microarray dataset obtained from the GEO profiles at National Centre for Biotechnology Information (NCBI) website. Cross validation of the generated rule sets shows 100% accuracy. Now to extract the hidden biological dependencies between responsible genes, Decision Tree is used at consecutive two stages of cancer development to identify the main culprit genes for cancer development from one stage to another and that may lead to the drug design. The analysis revealed that hybrid Rough- Decision Tree is able to extract hidden relationships among the various genes which play an important role in causing the disease and also able to provide a unique rule set for automated medical diagnosis. Moreover at the end, the functions of the identified genes are studied and validated from Gene Ontology website DAVID which clearly shows the direct or indirect relation of genes with the cancer. This study highlights the usefulness and efficiency of RST and Decision Tree in the disease diagnosis process and its potential use in inductive learning and as a valuable aid for building more biologically significant expert systems in medical sciences

DNA Diagnosis in Oncology

It is well established that cancers are caused by the accumulation of genomic and epigenomic alterations. However, molecular changes occuring in the early stages of cancer development or in precursor lesions remain to be poorly understood.The employment of molecular characterization of tumors prior to therapy has opened the door for personalized therapy of individual patients. Such genetic alterations that are utilized to predict response to therapy are considered predictive biomarkers. The development of high throughput technologies lead to the establishment of a relatively new field of cancer genomics. These genome-wide approaches are increasingly more important in cancer diagnostic, prognosis, and treatment. Here we briefly discuss cancer transcriptome, cancer genome and epigenome analyses approches and their clinical utility. Gene expression patterns in tumors (cancer transcriptome) are analyzed by expression microarrays who can serve as a diagnostic tool which facilitates distinguishing different cancer subtypes. Another high throughput technology that may revolutionize personalized cancer therapy, lead to the development of new genetic diagnostic tests and biomarkers is next generation sequencing (NGS). NGS has an ability to fully sequence large number of genes in a single test and simultaneously detect deletions, insertions, copy number alterations, translocations and exome-wide base substitutions in cancer-related genes. NGS methods analyses DNA methylation, detection of modified histones, mapping of transcription factor occupancy, and epigenetic regulators. DNA diagnosis often trumps clinical diagnosis in oncology. This will cause the shift toward testing minute amounts of DNA and specimens including needle biopsies, circulating tumor cells, and ciruclating cell-free DNA are likely to become clinically relevant. Thus, as NGS enters clinical testing it will impact clinical decisions and cancer outcomes. Finally, improved understanding of cancer biology will lead to further development of targeted therapies and the use of combinations of targeted approaches simultaneously, increasing treatment effectiveness and supporting highly individualized patient care.

119 Fork stalling at AT-rich sequences and failure of origin activation lead to chromosomal instability at fragile sites

Perturbed DNA replication in early stages of cancer development induces chromosomal instability preferentially at fragile sites. However, the molecular basis for this instability is unknown. Using DNA combing, we studied the replication dynamics along two common fragile sites on chromosome 16, FRA16C and FRA16D. We found in FRA16C that under normal growth conditions, the replication along the fragile region shows stress-like dynamics. Replication forks along the fragile site progress significantly slower than in the entire genome, and frequently stall at AT-rich sequences. Interestingly, under these conditions, most of the stalled forks are concentrated near the largest 3.3Kbs AT-rich sequence. Furthermore, the distance between origins along FRA16C are significantly shorter than in the entire genome. Together, these results indicate that even under normal growth conditions, replication along the fragile site is continuously perturbed leading to activation of additional origins to enable replication of the region. Under mild replication stress induced by aphidicolin, the replication rate at the FRA16C region is further slowed down and the frequency of fork stalling at AT-rich sequences is further increased. Strikingly, unlike the entire genome, additional origins are not activated suggesting that all potential origins in the FRA16C region are already activated under normal conditions. These results demonstrate the inability of FRA16C to compensate for replication stress leading to failure of normal replication completion (Ozeri-Galai et al., 2011). To analyse directly, the role of AT-rich sequences in fragile site expression we targeted the 3.3 Kbs long AT-rich sequence derived from FRA16C into a non-fragile genomic region using a homologous recombination-based system. The results of these experiments will be discussed. Replication dynamic analysis of another fragile site, FRA16D, which is enriched with AT-rich sequences, reveals for the first time that a combination of DNA sequences and abnormal origin activation underlies the molecular basis of replication stress sensitivity of a fragile site. Moreover, the results provide a novel mechanism underlying the replication delay and replication stress sensitivity of fragile sites and shed light on the basis of genomic instability during the early stages of cancer development.

MicroRNA Dysregulation in Gastric Cancer

Gastric carcinogenesis is a complex multistep process involving genetic dysregulation of proto-oncogenes and tumorsuppressor genes, and has recently entered the era of microRNAs (miRNAs), a class of small non-coding RNAs that posttranscriptionally regulate gene expression and control various cellular functions. MicroRNAs are small (approximately 22 nucleotides) non-coding RNAs that play fundamental roles in diverse biological and pathological processes, including cell proliferation, differentiation, apoptosis, and carcinogenesis. MicroRNAs have been revealed to be involved in various stages of cancer development, showing that abnormal miRNA expressions play critical roles in modulating expression of known oncogenes or tumor suppressor genes during cancer progression. Therefore, microRNAs can perform the function of onco-miRs or tumor-suppressor-miRs in gastric carcinogenesis. This review summarizes a current understanding of the connection between miRNAs and gastric cancer. Additionally, this paper outlines the applications of miRNAs in clinical practice, such as diagnosis, prognosis, detection, and therapy of gastric cancer.

Involvement of PTEN in TPA-mediated p53-activation in mouse skin epidermal JB6 cells

Our recent studies suggest a unique role of p53 during the early stage of cancer development. However, how p53 activation is regulated during TPA treatment remains elusive. We used murine skin epidermal JB6 promotion-sensitive (P+) and promotion resistant (P−) cells to observe differential expression of PTEN during TPA-induced p53 activation. Total PTEN expression was decreased in only P+ cells. Nuclear expression of PTEN increased and complex formation between PTEN and p53 occurred in P+ cells treated with TPA. Knocking down PTEN expression via siRNA inhibited TPA-induced Bax expression. Similar effects were seen with the p53 inhibitor, pifithrin-alpha. Cells that were transfected with siRNA to PTEN exhibited enhanced tumorigenicity. Our findings suggest PTEN mediates TPA-induced p53 activation. Structured summary of protein interactionsPTENandp53colocalizebyfluorescence microscopy(View interaction)PTENphysically interactswithp53byanti bait coimmunoprecipitation(View Interaction:1,2)PTENandp53colocalizebycosedimentation in solution(View interaction)

Consensus Micro RNAs Governing the Switch of Dormant Tumors to the Fast-Growing Angiogenic Phenotype

Tumor dormancy refers to a critical stage in cancer development in which tumor cells remain occult for a prolonged period of time until they eventually progress and become clinically apparent. We previously showed that the switch of dormant tumors to fast-growth is angiogenesis dependent and requires a stable transcriptional reprogramming in tumor cells. Considering microRNAs (miRs) as master regulators of transcriptome, we sought to investigate their role in the control of tumor dormancy. We report here the identification of a consensus set of 19 miRs that govern the phenotypic switch of human dormant breast carcinoma, glioblastoma, osteosarcoma, and liposarcoma tumors to fast-growth. Loss of expression of dormancy-associated miRs (DmiRs, 16/19) was the prevailing regulation pattern correlating with the switch of dormant tumors to fast-growth. The expression pattern of two DmiRs (miR-580 and 190) was confirmed to correlate with disease stage in human glioma specimens. Reconstitution of a single DmiR (miR-580, 588 or 190) led to phenotypic reversal of fast-growing angiogenic tumors towards prolonged tumor dormancy. Of note, 60% of angiogenic glioblastoma and 100% of angiogenic osteosarcoma over-expressing miR190 remained dormant during the entire observation period of ∼ 120 days. Next, the ability of DmiRs to regulate angiogenesis and dormancy-associated genes was evaluated. Transcriptional reprogramming of tumors via DmiR-580, 588 or 190 over-expression resulted in downregulation of pro-angiogenic factors such as TIMP-3, bFGF and TGFalpha. In addition, a G-CSF independent downregulation of Bv8 was found as a common target of all three DmiRs and correlated with decreased tumor recruitment of bone marrow-derived CD11b+ Gr-1+ myeloid cells. In contrast, antiangiogenic and dormancy promoting pathways such as EphA5 and Angiomotin were upregulated in DmiR over-expressing tumors. This work suggests novel means to reverse the malignant tumor phenotype into an asymptomatic dormant state and may provide promising targets for early detection or prevention of cancer.

Upregulation and inhibition of the nuclear translocation of Oct4 during multistep gastric carcinogenesis

Gastric cancer is the fourth most commonly diagnosed malignancy and the second leading cause of cancer-related mortality worldwide. Recent research suggests that tissue stem cells and the self renewal transcription factor, octamer-binding transcription factor4 (Oct4), could be involved in the development of certain tumors. The aim of this study was to investigate the expression pattern of Oct4 in normal human stomach and during multistep gastric carcinogenesis. Pyloric antral mucosal tissues were obtained from consenting individuals undergoing endoscopy (due to upper gastrointestinal symptoms) and gastrectomy (due to pyloric antral adenocarcinoma). Some tissue samples were processed to assemble an array of tissue sections representing multistep carcinogenesis and probed using anti-Oct4 antibodies and lectins specific for α-L-fucose or N-acetyl-D-glucosamine. Some tissue samples were processed for subcellular fractionation and western blot analysis using the same antibodies. The results revealed that Oct4-expressing cells were found in the proliferative cell compartment of the pit-gland units of microscopically normal gastric mucosal biopsies. Mucosal tissues with evidence of severe gastritis, metaplastic/dysplastic transformation and gastric cancer showed a significant increase in the expression of Oct4 (the labeled area increased from 2% in the control to 6 and 16% in the gastritis and cancerous tissues, respectively), suggesting a role for Oct4 in the early stages of cancer development. Furthermore, the data revealed an alteration in the subcellular distribution of Oct4, possibly due to the inhibition of cytoplasm-to-nucleus translocation during carcinogenesis. In conclusion, this study demonstrates an alteration in the expression pattern and nuclear translocation of Oct4 during gastric carcinogenesis and may be helpful in designing new modalities for the early detection and/or therapy of gastric cancer.

12 The Molecular Basis for Replication-Induced DNA Damage in Early Stages of Cancer Development

12 The Molecular Basis for Replication-Induced DNA Damage in Early Stages of Cancer Development

Isocitrate dehydrogenase 1 is downregulated during early skin tumorigenesis which can be inhibited by overexpression of manganese superoxide dismutase

Isocitrate dehydrogenase 1 (IDH1), a cytosolic enzyme that converts isocitrate to alpha-ketoglutarate, has been shown to be dysregulated during tumorigenesis. However, at what stage of cancer development IDH1 is dysregulated and how IDH1 may affect cell transformation and tumor promotion during early stages of cancer development are unclear. We used a skin cell transformation model and mouse skin epidermal tissues to study the role of IDH1 in early skin tumorigenesis. Our studies demonstrate that both the tumor promoter TPA and UVC irradiation decreased expression and activity levels of IDH1, not IDH2, in the tumor promotable JB6 P+ cell model. Skin epidermal tissues treated with dimethylbenz[α]anthracene/TPA also showed decreases in IDH1 expression and activity. In non-promotable JB6 P-cells, IDH1 was upregulated upon TPA treatment, whereas IDH2 was maintained at similar levels with TPA treatment. Interestingly, IDH1 knockdown enhanced, whereas IDH1 overexpression suppressed, TPA-induced cell transformation. Finally, manganese superoxide dismutase overexpression suppressed tumor promoter induced decreases in IDH1 expression and mitochondrial respiration, while intracellular alpha-ketoglutarate levels were unchanged. These results suggest that decreased IDH1 expression in early stage skin tumorigenesis is highly correlated with tumor promotion. In addition, oxidative stress might contribute to IDH1 inactivation, because manganese superoxide dismutase, a mitochondrial antioxidant enzyme, blocked decreases in IDH1 expression and activity.