Pancreatic Adenosquamous Carcinoma Case Study 1

Superior Essays
Pancreatic adenosquamous carcinoma (ASC) is a rare and aggressive tumor that has worse prognosis and higher metastatic potential than pancreatic adenocarcinoma(Kardon, Thompson et al. 2001, Boyd, Benarroch-Gampel et al. 2012). A major hindrance towards development of therapies against pancreatic ASC is that no unique molecular signature has been identified for this class of pancreatic tumors. Although KRAS and TP53 are altered in pancreatic ASC (Brody, Costantino et al. 2009), the same genes are also abnormal in other forms of pancreatic cancer. The molecular mechanism that promotes pancreatic adenosquamous carcinoma is unknown. The main objective of this proposal is to identify new driver genes for pancreatic adenosquamous carcinoma that can …show more content…
To identify how UPF1 mutations alter gene expression, I will utilize CRISPR based genome editing (Doudna and Charpentier 2014) to generate human cell lines that contain UPF1 mutations identified in pancreatic ASC. To examine the differentially expressed candidates in pancreatic ASC, I will isolate total RNA from UPF1 mutant cell lines and perform RNA sequencing. To establish whether the upregulated genes are direct targets of NMD, I will compare this data with RNA isolated from cells where UPF1 is downregulated using short hairpin RNAs. To validate the top targets from RNA sequencing experiments, I will perform qPCR and northern blotting. To examine whether these candidates are also altered in pancreatic ASC, I will analyze the RNA levels of top targets in pancreatic ASC samples that contain UPF1 mutations.
To determine the function of the identified candidates from high-throughput sequencing, I will perform overexpression and knockdown experiments in primary pancreatic cell lines to examine for corresponding gain or loss in proliferation. In addition, I will determine whether overexpression of the candidates transforms primary pancreatic cells. In summary, results from this aim will elucidate the mechanism by which UPF1 mutations promote pancreatic
…show more content…
Since Upf1 knockout is embryonic lethal but Upf1 heterozygotes are normal (Medghalchi, Frischmeyer et al. 2001), I will generate conditional Upf1 mutants. Previous studies have shown that majority of UPF1 mutations identified in pancreatic ASC promotes skipping of exons 10 and exon11 that cause a truncation in the essential RNA helicase domain. Mutations in the RNA helicase domain of UPF1 has been shown to be dominant negative(Sun, Perlick et al. 1998). To model this exon-skipping event, I will generate a targeting construct that will place the loxP sites on the introns flanking exon 10 and exon11 upon homologous recombination in ES cells. To control the expression of the floxed Upf1 allele, I will place a lox-stop-lox cassette in the first intron of Upf1 that causes transcription termination in the absence of Cre-recombinase. To restrict expression of Upf1 mutants to the pancreas, I will cross the Upf1 floxed mice with PDX1-CRE expressing mice(Hingorani, Petricoin et al. 2003).
To determine whether Upf1 mutants promote pancreatic ASC, I will monitor 20 wild type controls and 20 mutant animals. Survival curves, tumor incidence, and pancreatic tumor types arising in wild type and mutant animals will be compared. In conclusion, these experiments will demonstrate whether UPF1 mutations are essential for pathogenesis of cancer in mice and potentially in

Related Documents

  • Improved Essays

    Lung Cancer Essay

    • 1525 Words
    • 7 Pages

    Carcinogens bring about mutation in the genes that induce development of cancer. About 30 percent of lung adenocarcinomas are brought about by a mutation in the K-ras proto-oncogene. It has also been reported that epigenetic alterations that happen on genes may lead to inactivation of tumor suppressor genes. This, therefore, implies that proliferation of cells will be uncontrollable leading to lung cancer. The epidermal growth factor receptor (which functions to regulate apoptosis, cell proliferation and tumor invasion) undergoes mutation.…

    • 1525 Words
    • 7 Pages
    Improved Essays
  • Improved Essays

    CRISPR Essay

    • 760 Words
    • 4 Pages

    The possibility of being capable of changing the human genome to prevent or treat devastating illnesses and serious inherited diseases has always being in the mind of many people and scientists. However, two important events may contribute to make this idea a reality in the near future. One is the decoding of human genome. This has allow scientists the opportunity to comprehend how the genetic information controls the growth, configuration and function of the human body; and at the same time, to understand how variations within our DNA sequence cause diseases.…

    • 760 Words
    • 4 Pages
    Improved Essays
  • Superior Essays

    Pancreatic Cancer Essay

    • 1222 Words
    • 5 Pages

    Pancreatic Cancer In the U.S an estimated 46,420 will be diagnosed with pancreatic cancer and over 39,590 will die from this disease. For stage IA there’s only a 14% chance of survival and for stage IV there’s only a 1% chance. Not many people want to know these statistics, because they might lose hope if they are fighting the disease; for those that would like to gain an understanding it should be made clear. Knowing this, it’s best to learn the facts of how this cancer can be prevented and diagnosed.…

    • 1222 Words
    • 5 Pages
    Superior Essays
  • Superior Essays

    Fruit Fly Lab Report

    • 1243 Words
    • 5 Pages

    Name: Enrique Vazquez PS ID #: 1259993 BIOL 3311 Fall 2014 Date: October 2, 2014 Gene: Bar TA Instructor Name: Fahmi Mesmar Lab Section: 16258 Writing Assignment 3: Eye morphology of Drosophila melanogaster; Bar mutation effects on ommatidial development of the compound eye. Introduction: Drosophila melanogaster have been used for centuries as model organisms due to their relatively small size as well as their inexpensive and simple diet. Fruit flies also have to ability to reproduce in large amounts considering their relatively short life cycles.…

    • 1243 Words
    • 5 Pages
    Superior Essays
  • Improved Essays

    Lafora disease is a rare, inevitably fatal, autosomal recessive, progressive myoclonus epilepsy. Being autosomal recessive, the disease is most commonly seen in communities that practice consanguinity. The onset of the disease occurs within the first two decades of life, with an average life span of 10 years following diagnosis (Monaghan & Delanty, 2010). Lafora disease is characterized by myoclonic and tonic-clonic seizures and neurological dysfunction, such as dementia (Monaghan & Delanty, 2010). The hallmark of Lafora disease is the accumulation of inclusions, called Lafora bodies, in the cytoplasm of neuronal somas, the heart, liver, muscles, and other tissues (Roach, 2015).…

    • 765 Words
    • 4 Pages
    Improved Essays
  • Improved Essays

    Determination of GAPC Gene Sequence for Brassica Oleracea var. acephala and Brassica Oleracea var. capitate INTRODUCTION GAPDH is an enzyme that is encoded by the GAPC gene and catalyzes glycolysis’ sixth step which is responsible for energy metabolism (Bio Rad 2017). Aside from its role in glycolysis, GAPDH is also known for its evolutionary significance specifically that it has roles in cellular function like nuclear RNA export and DNA replication and repair (Tatton et al. 2001). GAPDH has been found in apoptosis, neurodegenerative disease, and viral pathogens which is part of its biomedical significance (Tatton et al. 2001).…

    • 650 Words
    • 3 Pages
    Improved Essays
  • Improved Essays

    Soto's Syndrome Analysis

    • 412 Words
    • 2 Pages

    Sotos syndrome is a disease that causes a child to grow unusually rapidly, especially in the cephalic region, resulting in abnormal growth of the brain and a corresponding array of developmental disabilities (Türkmen et al., 2015; Faravelli, 2005; McKusick, 2015). This disorder is an autosomal dominant disorder (McKusick, 2015) and has been found to be a result of mutations in the NSD1 gene (Türkmen et al., 2015; Faravelli, 2005; McKusick, 2015), on the chromosome 5q35 (Faravelli, 2005; McKusick, 2015). There are over 100 mutations that have been found in the NSD1 gene, with all of them occurring de novo and at no specific spot in the gene (Faravelli, 2005). NSD1 contains 8088 base pairs, 23 exons, and translates into a protein that is 2696 amino acids long (Faravelli, 2005). This gene, along with the rest of the NSD family, have been found to be strongly linked to cell growth and differentiation (Faravelli, 2005).…

    • 412 Words
    • 2 Pages
    Improved Essays
  • Improved Essays

    Step 2) Production of CRISPR RNA – CRISPR repeats and…

    • 647 Words
    • 3 Pages
    Improved Essays
  • Superior Essays

    Hereditary Cancer Cancer is one of the most unknown subjects in the medical world. When it comes, why it comes, and how to effectively contain it is still being studied. Treatments have been available to lessen the cancer, or in some cases, take it away. The medical community has come so far in our knowledge of cancer. Splitting cancer into two specific groups of somatic cancer and hereditary cancer, researchers are getting more information.…

    • 1357 Words
    • 6 Pages
    Superior Essays
  • Improved Essays

    By being able to pinpoint one of the genes leading to this particular cancer, scientists can better understand where and why this mutation…

    • 1078 Words
    • 5 Pages
    Improved Essays
  • Great Essays

    For example, diets high in red and processed meats, cooking meats at high temperatures creates chemicals linked to colon cancer, lack of physical activity, smoking, and heavy alcohol use. People with type II diabetes may also have an increased risk of developing colon cancer as well as people with an history of inflammatory bowel disease (Cancer Treatment Centers of America, 2001). People can inherit genes that elevate Colon Cancer risks as well as having shared environmental factors. The most common inherited syndromes corresponding with colorectal cancer are Familial Adenomatous Polyposis (FAP) and Hereditary non-polyposis Colorectal Cancer (HNPCC) or Lynch Syndrome (Mayo Clinic 2013). A change in the APC gene causes FAP.…

    • 1042 Words
    • 5 Pages
    Great Essays
  • Improved Essays

    Folliculin Case Study

    • 884 Words
    • 4 Pages

    Aim 1: Determine the amino acid sensing specificity of folliculin. Since folliculin’s has novel molecular function in amino acid sensing I plan to further understand whether FLCN specifically senses a certain amino acid and if this level of detection is enough to elicit an mTORC1 response. 1.1. Determine if folliculin’s novel molecular function in amino acid sensing is triggered by glutamine. mTORC1 activation by amino acids requires FLCN, whereby its lysosomal residence during starvation interacts with inactive Rag complex and exerts GAP activity upon RagC/D during amino acid re-stimulation.…

    • 884 Words
    • 4 Pages
    Improved Essays
  • Improved Essays

    Genome targeting and editing has provided the ability to better understand and determine the function of genes in many model organisms. Genomes are composed of genes which are the basic unit of heredity that determine the traits organisms express. With the ability to target specific genes and edit them, genes and their locations in the genome can be studied and determine their significance and function, in addition to gaining an understanding to human disease and possible routes to eliminate them. The most effective methods of genome editing are those of the zinc-finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), and the clustered regulatory interspaced short palindromic repeats/CRISPR-associated systems (CRISPR/Cas).…

    • 862 Words
    • 4 Pages
    Improved Essays
  • Superior Essays

    cerevisiae and drosophila (Rothstein 1991; S. Rong 2002). Especially in yeast cells, HR is commonly used method for gene targeting due to high efficiency. Yeast cells can grow as haploid/diploid forms; it provides specific advantage to study lethality-gene relationship and yeast mating type switch stimulates the efficient homologous recombination machinery. Target gene can be disrupted by using plasmid containing nonfunctional copy of gene, selectable marker flanked by short ~40 nucleotides homologous to target gene [5]. Basically, nonfunctional copy of gene is cloned into plasmid vector containing selectable marker.…

    • 1692 Words
    • 7 Pages
    Superior Essays
  • Improved Essays

    I was keenly aware of the importance of Genetics in health care early in my time studying Medicine and later have gained profound knowledge of it as well as Cell Biology and Molecular Biology through in-depth Biomedical studies. However, it is during my current study on the genetic disorder Neurofibromatosis type 1 (NF1) when my passion for the Laboratory Genetics and Genomics was lighted. My passion for the Laboratory Genetics and Genomics was lighted while I am conducting current studies on the genetic disorder Neurofibromatosis type 1 (NF1). However, it has long been deeply rooted since I was keenly aware of the importance of Genetics in health care early in my time studying Medicine and have gained profound knowledge of it as well as Cell…

    • 726 Words
    • 3 Pages
    Improved Essays