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1 Tumor Markers Currently Utilized in Cancer Care Gaetano Tumor markers currently utilized in cancer care Gaetano Romano (gromano at temple dot edu) Department of Biology, College of Science and Technology, Temple University, Bio-Life Science Bldg. Suite 456, 1900 N 12th Street, Philadelphia, PA 19122, U.S.A. DOI http://dx.doi.org/10.13070/mm.en.5.1456 Date last modified : 2015-10-02; original version : 2015-09-25 Cite as MATER METHODS 2015;5:1456 Abstract A review of tumor markers that are currently used in cancer care. Introduction Tumor markers are products that may derive from malignant cells and/or other cells of the organism in response to the onset of cancer [1-5]. Their production may also be induced by noncancerous benign tumors [1-5]. Some tumor markers can be detected in malignant tissues obtained from biopsies [6-19], whereas others can be analyzed in the blood, bone marrow, urine, or other body fluids [20-25]. Sometimes, tumor markers may also be observed in cancer-free subjects, but in much lower doses than oncological patients. In addition, relatively high levels of a certain tumor marker might develop from various non-malignant pathological conditions, such as liver diseases, inflammations, kidney-related dysfunctions, infections and hematological disorders. On these grounds, high levels of a certain tumor marker in the blood, or in other body fluids might indicate the presence of a malignancy. However, per se, this finding is not sufficient to substantiate the diagnosis of a cancer. For this reason, the analysis of tumor markers in the blood, or other fluids must be combined with the analysis of biopsies, or other tests in order to confirm the diagnosis. The detection of tumor markers can be used for a wide variety of malignancies and for a number of applications, which may comprise diagnosis, follow-up the clinical course of the disease, optimize the efficacy of the treatment, assess the response to therapy and monitor the recurrence of the disease [1-5]. The National Academy of Clinical Biochemistry and The American Society of Clinical Oncology (ASCO) provide the guidelines for the proper utilization of tumor markers in cancer care, which must be approved by the Food and Drug Administration (FDA), in order to be used in the clinical setting of the United States of America (U.S.A.). This article describes the methods that are adopted for the detection of tumor markers that are currently used in cancer care, along with new strategies that aim at developing personalized medicine programs for the treatment of malignancies. Tumor marker Type of tumor/tumors Application 21-Gene signature (Oncotype Assess risk of tumor Breast cancer. DX). recurrence. 70-Gene signature Assess risk of tumor Breast cancer. (Mammaprint). recurrence. Anaplastic lymphoma kinase Non-small cell lung cancer and Diagnosis and determine (ALK) (mutations). anaplastic large cell lymphoma. type of treatment. Diagnosis and determine Melanoma, colorectal cancer and BRAF (mutations). type of treatment in patients thyroid cancer. with melanoma. 1 Cancers of the breast, head and Epidermal growth factor Predict outcomes and neck, non-small cell lung, receptor (EGFR), or HER1 determine type of treatment. pancreas and colon. HER2, or HER2/neu, erbB-2, Breast cancer, stomach cancer and Predict outcomes and EGFR2 esophageal cancer. determine type of treatment. Breast cancer and gynecologic Hormone receptor (estrogen malignancies, such as endometrial Predict outcomes and and progesterone) stromal sarcomas and endometrial determine type of treatment. cancers. Gastrointestinal stromal tumor and Diagnosis and determine KIT mucosal melanoma. type of treatment. Advanced colorectal cancer and KRAS (mutations) Determine type of treatment. lung cancer. Diagnosis. Blood tests allow for the follow-up of the S-100 Melanoma. clinical course of the disease. Urokinase plasminogen To assess the aggressiveness activator (uPA) and of the malignancy and Breast cancer. plasminogen activator determine the type of inhibitor (PAI-1). treatment. Table 1. List of tumor markers that are detected in malignant tissues, other than the blood. Tumor markers detected in malignant tissues The detection of these tumor markers requires the removal of a biopsy from the patient [6-19]. In certain cases, biopsy-derived tumor markers are utilized just once, in order to confirm a diagnosis. The list of tumor markers that are analyzed in malignant tissues is summarized in Table 1. 21-Gene signature (Oncotype DX) This set of markers is utilized to assess the risk of the reappearance of the tumor in patients with breast cancer [9,26-28]. The 21-gene signature comprises genes that are related to the estrogen receptor (ESR1, PGR, BCL2, SCUBE2), HER2 (HER2 and GRB7), cell proliferation (Ki67, STK15, survivin, cyclin B1, MYBL2), cellular invasion (stromelysin3, cathepsin L2), macrophage marker CD68, anti-apoptotic genes (BAG1 and GSTM1) and five housekeeping genes that are used as reference (-actin, GAPDH, RPLPO, GUS and TFRC) [9, 26-28]. Oncotype DX is a gene-expression profiling that requires RNA samples derived from paraffin- embedded tumor biopsies and was introduced for the first time in the U.S.A. market in 2004 (Genomic Health Inc., Redwood City, CA) [29]. 70-Gene signature (MammaPrint) This set of markers is utilized to assess the risk of the recurrence of the tumor in patients with breast cancer (for a review see references 31-35). In contrast to the 21-Gene signature (Oncotype DX), this assay needs a preparation of fresh tissues in a solution designed to preserve the integrity of RNA molecules [31]. The MammaPrint 70-gene signature test is provided by Agendia Inc. (Irvine, CA). Anaplastic lymphoma kinase (ALK) The ALK gene is analyzed for the presence of mutations [33-35]. This tumor marker may provide useful information both for the prognosis and to determine the type of treatment for non-small cell lung cancer (NSCLC) and anaplastic large cell lymphoma (ALCL) [33-35]. If mutations are 2 present, the patient can be treated with crizotinib (or Xalkori), which is a protein kinase inhibitor that targets the aberrant ALK kinase [33, 35]. However, crizotinib interferes also with some other protein kinases [36, 37]. ALK mutation assay analyzes tissues by means of various techniques, such as fluorescent in situ hybridization (FISH), immunohistochemistry (IHC) and polymerase chain reaction (PCR) [35]. FDA approved in 2011 the ALK Break Apart FISH Probe Kit (Abbot Vysis, Schaumburg, IL). This procedure requires unstained tissues, which are hybridized overnight with the probe and then analyzed by fluorescence microscopy [35]. IHC-based techiniques work well for the ALK detection in ALCL, However, IHC-based ALK detection is not efficient in NSCLC biopsies, because of lower levels of ALK protein. IHC detection of ALK in ALCL utilizes the following antibodies: ZAL4 (Invitrogen, Carlsbad, CA), 5A4 (Novocastra, Newcastle, UK) and D5F3 (Cell Signaling Technology, Denvers, MA). Lastly, PCR-derived techniques are able to detect the presence of ALK in NSCLC with various protocols, which comprise reverse-transcriptase multiplexed PCR and RT-PCR [35]. BRAF Mutations of the BRAF gene may be present in melanoma [11, 38, 39], colorectal cancer [40, 41] and thyroid cancer [42, 43]. This tumor marker is used for the diagnosis and to determine the type of treatment in patients with melanoma [11, 38, 39]. The so-called BRAF V600E mutation may be detected in approximately 50% of cases of melanoma [44]. Patients with advanced melanoma can be treated with vemurafenib (or Zelboraf), if mutations in the BRAF gene are found [45, 46]. Vemurafenib has the ability to target the abnormal BRAF protein [45-47]. Various assays are currently available for the detection of the BRAF V600E mutation in formalin-fixed paraffin- embedded (FFPE) tumor tissues, which may also be mounted on slides. Such assays comprise techniques based on pyrosequencing (Laboratory Corporation of America, Burlington, NC), dideoxy sequencing (Quest Diagnostics Inc., Madison, NJ; Vanderbilt Pathology Laboratory Service, Nashville, TN) and PCR either with or without fluorescence monitoring (ARUP Laboratories, Salt Lake City, UT; Mayo Medical Laboratories, Rochester, MN; UNC Health Care McLendon Clinical Laboratories, Chapel Hill, NC). Epidermal growth factor receptor (EGFR) EGFR is also termed HER1 [48, 49]. The overexpression of this cell surface receptor may be indicative of poor clinical outcomes, as cancerous tissues growth fast, malignant cells spread rapidly and the tumor tends to be more resilient to the treatment [48-52]. EGFR may be used to predict outcomes and to determine the type of treatment for cancers of the breast [53-56], head and neck [57-59], non-small cell lung [34, 51, 60, 61], pancreas [62, 63] and colon [64-66]. Tumor tissues must be prepared in formalin-fixed paraffin-embedded (FFPE) samples. A real- time PCR test analyses mutations in exons 18, 19, 20 and 21 of the EGFR gene, which may be present in DNA obtained from FFPE human non-small cell lung cancer (NSCLC) tumor tissues (Roche Diagnostics, cobas® EGFR Mutation Test, Indianapolis, IN). Another PCR-based method utilizes the therascreen® EGFR RGQ PCR Kit - P120022 kit (Qiagen Manchester Ltd, Manchester, U.K.) for the analysis of FFPE human NSCLC tumor tissue-derived DNA. A qualitative IHC-based system utilizes the EGFR pharmDxTM kit to monitor EGFR expression levels in normal and malignant tissues of the colon and of head and neck squamous carcinoma, which must be fixed for histological analysis (Dako North America, Inc., Carpinteria, CA). A chromogenic in situ hybridization CISH protocol was developed for the analysis of FFPE breast cancer tissues (Zymed Laboratories, Inc., New York. NY). FFPE breast cancer tissue may also be analyzed by IHC, utilizing a monoclonal antibody to EGFR (Clone 31G7, Zymed Laboratories, Inc., New York. NY). HER2 HER2 is also known as HER2/neu, erbB-2, or EGFR2 [67]. This cellular receptor may contribute to the growth and/or dissemination of certain types of tumors [14, 67, 68].
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