The long Non-coding RNA Orchestrator of Cancer Axis of EvilInsights into the Multiple Modes of Action of the H19 Gene

ncreasing evidence has indicated that the non-coding RNA molecules play central roles in almost all biological processes and many pathological conditions including carcinogenesis. This review focuses on the pathological tumorigenic role of the first discovered long non-coding RNA gene called H19 and its pivotal contribution to the cancer axis of evil. H19 RNA utilizes a variety of mechanisms to perform its pathological function. Some key unanswered questions are presented by the end. Understanding the H19 RNA mechanisms of action will shed light into the class of long non-coding RNA which contains contains thousands of members mostly with unknown functionn and will help in delineating the pathological role played by at least some of the

Development of a PCR /RLB Test for meat source authentication in processed meat and meat products

Concern over food authenticity has increased as a result of an increase in the consumption of processed foods containing meat or animal products. This raises a number of issues where the presence of pork in such foods is considered unacceptable in most Muslim and Jewish communities around the world. It also applied to the prohibition of beef consumption among Hindus. In order to ensure the absence of unwished meat products or mixing of meats from different sources in processed foods, a specific and sensitive test is essential. For this purpose we developed a molecular test based on DNA amplification by polymerase chain reaction (PCR) of the cytochrome b gene followed by reverse line blot analysis (RLB). Using this method many samples may be treated simultaneously and meat origins can easily be detected from processed foods or foods containing mixed meat sources; also, added pork components such as fat may be identified by this methodology. The PCR/RLB method is considered to be a sensitive and specific technique; it can detect one nucleotide change within the PCR-amplified DNA segment.This study was supported by the Arab Science and Technology Foundation, grant number BT072239

Strategy for DNA extraction and detection frominsect pests in stored home grain samples

Stored grains are subjected to infestations with more than 60 species of insects, that responsible for millions of dollars loss and cause several health problems including allergies and gastrointestinal disorders. Traditional detection techniques are laborious, expensive and not sensitive to detect insect contamination at the egg and larvae stages. Therefore, alternative methods are needed for rapid and sensitive detection. One obvious approach is to develop a molecular approach utilizing genetic information of the potential insect species that infest grains for amplification of specific target gene fragment utilizing polymerase chain reaction [PCR]. In the present study, a number of known infested grain samples were used in standardizing a method to isolate larvae and adult insects that were based on centrifugation washing method and a filtration washing method. The isolated insects were subjected to DNA extraction and PCR amplification of defined regions of cytochrome oxidase I (COI) gene followed by sequencing to identify the different pest species. For PCR amplification new primers were designed and for this purpose the obtained COI sequences from different insects were aligned to design two sets of primers (named: COI-PCR4 and COI-PCR5) specific for the indicated insect mitochondrial COI gene. The designed primers were tested for their specificity and sensitivity. The suitability of PCR primers and DNA extraction methods were evaluated on eleven samples of commercial grains utilizing each primer set with the two extraction methods

The H19 Non-Coding RNA Is Essential for Human Tumor Growth

Background. Mutations and epigenetic aberrant signaling of growth factors pathways contribute to carcinogenesis. Recent studies reveal that non-coding RNAs are controllers of gene expression. H19 is an imprinted gene that demonstrates maternal monoallelic expression without a protein product; although its expression is shut off in most tissues postnatally, it is reactivated during adult tissue regeneration and tumorigenesis. Moreover, H19 is highly expressed in liver metastasis derived from a range of carcinomas. The objective of this study is to explore the role of H19 in carcinogenesis, and to determine its identification as an anti-tumor target. Methodology / Principle Findings. By controlling oxygen pressure during tumor cell growth and H19 expression levels, we investigated the role of H19 expression in vitro and in vivo in hepatocellular (HCC) and bladder carcinoma. Hypoxia upregulates the level of H19 RNA. Ablations of tumorigenicity of HCC and bladder carcinomas in vivo are seen by H19 knockdown which also significantly abrogates anchorage-independent growth after hypoxia recovery, while ectopic H19 expression enhances tumorigenic potential of carcinoma cells in vivo. Knocking-down H19 message in hypoxic stress severely diminishes p57 kip2 induction. We identified a number of potential downstream targets of H19 RNA, including angiogenin and FGF18. Conclusions. H19 RNA harbors pro-tumorigenic properties, thus the H19 gene behaves as an oncogene and may serve as a potential new target for anti-tumor therapy

H19-Promoter-Targeted Therapy Combined with Gemcitabine in the Treatment of Pancreatic Cancer

Pancreatic cancer is the eighth cancer leading cause of cancer-related death in the world and has a 5-year survival rate of 1–4% only. Gemcitabine is a first line agent for advanced pancreatic therapy; however, its efficacy is limited by its poor intracellular metabolism and chemoresistance. Studies have been conducted in an effort to improve gemcitabine treatment results by adding other chemotherapeutic agents, but none of them showed any significant advantage over gemcitabine monotherapy. We found that 85% of human pancreatic tumors analyzed by in situ hybridization analyses showed moderated to strong expression of the H19 gene. We designed a preclinical study combining gemcitabine treatment and a DNA-based therapy for pancreatic cancer using a non viral vector BC-819 (also known as DTA-H19), expressing the diphtheria toxin A chain under the control of the H19 gene regulatory sequences. The experiments conducted either in an orthotopic and heterotopic pancreatic carcinoma animal model showed better antitumor activity following the sequential administration of the vector BC-819 and gemcitabine as compared to the effect of each of them alone. The results presented in the current study indicate that treatment with BC-819 in combination with gemcitabine might be a viable new therapeutic option for patients with advanced pancreatic cancer

Treatment of ovarian cancer ascites by intra-peritoneal injection of diphtheria toxin A chain-H19 vector: a case report

<p>Abstract</p> <p>Introduction</p> <p>Ovarian cancer ascitic fluid, which contains malignant cells, is usually present in women with an advanced stage disease. There are currently no effective therapies for the treatment of ovarian cancer ascitic fluid. We developed a new therapeutic strategy to target expression of the diphtheria toxin fragment A gene in ovarian tumor cells under the control of H19 regulatory sequences.</p> <p>Case presentation</p> <p>A 64-year-old Caucasian woman was diagnosed with a stage IIIc epithelial ovarian cancer. She suffered from progressive disease, accumulation of malignant ascites that needed to be drained weekly, abdominal pain, vomiting, anorexia and severe weakness. Infusion of the diphtheria toxin A chain-H19 plasmid into the peritoneum of our patient resulted in complete resolution of the ascites with minimum adverse events.</p> <p>Conclusion</p> <p>On the basis of this preliminary experience, we are currently conducting an extensive Phase I study on a larger number of patients in order to assess the safety and preliminary efficacy of this novel patient-oriented treatment approach.</p

NAF-1 and mitoNEET are central to human breast cancer proliferation by maintaining mitochondrial homeostasis and promoting tumor growth

Mitochondria are emerging as important players in the transformation process of cells, maintaining the biosynthetic and energetic capacities of cancer cells and serving as one of the primary sites of apoptosis and autophagy regulation. Although several avenues of cancer therapy have focused on mitochondria, progress in developing mitochondria-targeting anticancer drugs nonetheless has been slow, owing to the limited number of known mitochondrial target proteins that link metabolism with autophagy or cell death. Recent studies have demonstrated that two members of the newly discovered family of NEET proteins, NAF-1 (CISD2) and mitoNEET (mNT; CISD1), could play such a role in cancer cells. NAF-1 was shown to be a key player in regulating autophagy, and mNT was proposed to mediate iron and reactive oxygen homeostasis in mitochondria. Here we show that the protein levels of NAF-1 and mNT are elevated in human epithelial breast cancer cells, and that suppressing the level of these proteins using shRNA results in significantly reduced cell proliferation and tumor growth, decreased mitochondrial performance, uncontrolled accumulation of iron and reactive oxygen in mitochondria, and activation of autophagy. Our findings highlight NEET proteins as promising mitochondrial targets for cancer therapy

Development of targeted therapy for ovarian cancer mediated by a plasmid expressing diphtheria toxin under the control of H19 regulatory sequences

<p>Abstract</p> <p>Background</p> <p>Ovarian cancer ascites fluid (OCAF), contains malignant cells, is usually present in women with an advanced stage disease and currently has no effective therapy. Hence, we developed a new therapy strategy to target the expression of diphtheria toxin gene under the control of H19 regulatory sequences in ovarian tumor cells. H19 RNA is present at high levels in human cancer tissues (including ovarian cancer), while existing at a nearly undetectable level in the surrounding normal tissue.</p> <p>Methods</p> <p>H19 gene expression was tested in cells from OCAF by the in-situ hybridization technique (ISH) using an H19 RNA probe. The therapeutic potential of the toxin vector DTA-H19 was tested in ovarian carcinoma cell lines and in a heterotopic animal model for ovarian cancer.</p> <p>Results</p> <p>H19 RNA was detected in 90% of patients with OCAF as determined by ISH. Intratumoral injection of DTA-H19 into ectopically developed tumors caused 40% inhibition of tumor growth.</p> <p>Conclusion</p> <p>These observations may be the first step towards a major breakthrough in the treatment of human OCAF, while the effect in solid tumors required further investigation. It should enable us to identify likely non-responders in advance, and to treat patients who are resistant to all known therapies, thereby avoiding treatment failure.</p