Molecular Biology - 02 Clinical Applications

MOLECULAR BIOLOGY AND DIAGNOSTICS CLINICAL APPLICATIONS OF MOLECULAR BIOLOGY • OUTLINE • • Diagnosis of Disease o Introduction o Cancer o Inherited Human Diseases o Infectious Diseases • Role of Molecular Biology in • Forensic Science o o Introduction o Difference of Forensic Testing and Clinical Laboratory Testing • o DNA Typing and the Characteristics o DNA Typing Systems and their Characteristics • DNA Fingerprinting o Introduction o Steps on How DNA Fingerprint is Produced Gene Mapping o Introduction o How do Researchers Create a Genetic Map? o Uses of Gene Mapping Probability and DNA Testing Introduction o Types of Probabilities o Rules in Probability o Application: Paternity Testing Genetic Screening o What is Genetic Screening? o Types of Genetic Screening o Screening Tests o How is Screening Done? o Advantages and Disadvantages of Genetic Screening • TYPES OF TUMORS • • LESSON 1: DIAGNOSIS OF DISEASE INTRODUCTION • Cancer is uncontrolled growth of abnormal cells that’s spread to other organs of the body and 100 to 350 in every 350,000 people dies every year. Cancer is non-infectious but an outcome of the cell failure mechanism to produce a normal cell. Infectious disease such as Human Papilloma virus, Hepatitis C and Cytomegalovirus are detected using Molecular method. • • CANCER • • • • The losses of cellular regulation that give rise to most or all cases of cancer result from genetic damage that is often caused by tumor-promoting chemicals, hormones, and sometimes viruses. Mutations in three broad classes of genes have been implicated in the onset of cancer. Repeated process of Mutation, competition and natural selection operating within the population of somatic cells can cause matters to go from bad to worse. Mutations in three broad classes of genes have been implicated in the onset of cancer. o Proto oncogenes normally promote cell growth; mutations change them into oncogenes whose products are excessively active in growth promotion. Oncogenic mutations usually result in either increased gene expression or production of a hyperactive gene product. o Tumor-suppressor genes normally restrain growth, so mutations that inactivate them allow inappropriate cell division. o Genome maintenance gene are involved in maintaining the genome’s integrity. When these genes are inactivated, cells acquire additional genetic changes at an increased rate including mutations that cause the deregulation of cell growth and proliferation and lead to cancer. Many of the genes in these three classes encode proteins that help regulate cell proliferation (example: entry into and progression through the cell cycle) or cell death by apoptosis; others encode proteins that participate in repairing damaged DNA. Cancer Cells Bypass Normal Proliferation Controls and Colonize Other Tissues : Cancer cells are defined by two heritable properties: o Reproduce in defiance of the normal restraints on cell growth and division. o Invade and colonize territories normally reserved for other cells. It is the combination of these properties that makes cancers particularly dangerous. The combination of these properties that makes cancers particularly dangerous. An abnormal cell that grows (increases in mass) and proliferates (divides) out of control will give rise to a tumor, or neoplasm-new growth. • Benign- neoplastic cells have not yet become invasive. For most types of such neoplasms, removing or destroying the mass locally usually achieves a complete cure. Malignant or true cancer- cells have acquired the ability to invade surrounding tissue. Invasiveness is an essential characteristic of cancer cells. It allows them to break loose, enter blood or lymphatic vessels, and form secondary tumors called metastases at other sites in the body. In general, the more widely a cancer spreads, the harder it becomes to eradicate. It is generally metastases that kill the cancer patient. Classification of Cancer in Malignant Tumors: o Carcinomas - are cancers arising from epithelial cells, and they are by far the most common cancers in humans. They account for about 80% of cases, perhaps because most of the cell proliferation in adults occurs in epithelia. o Sarcomas arise from connective tissue or muscle cells. Benign Tumors: o Adenoma - is a benign epithelial tumor with a glandular organization. o Chondroma and a chondrosarcoma benign and malignant tumors of cartilage. o Melanoma derived from a pigment cell in the skin, will often (but not always) continue to make pigment granules. METASTASIS • Malignant tumors typically give rise to metastases, making the cancer hard to eradicate. Shown in this fusion image is a whole-body scan of a patient with metastatic nonHodgkin’s lymphoma (NHL). The background image of the body’s tissues was obtained by CT (computed x-ray tomography) scanning. Overlaid on this image, a PET (positron emission tomography) scan reveals the tumor tissue (yellow), detected by its unusually high uptake of radioactively labeled fluorodeoxyglucose (FDG). High FDG uptake occurs in cells with unusually active glucose uptake and metabolism, which is a characteristic of cancer cells. The yellow spots in the abdominal region reveal multiple metastases. (Courtesy of S. Gambhir.) Diseases Tay-Sachs disease Molecular and Incidence Cellular Defect Defective hexosaminidase 1/1000 eastern enzyme leads to European Jews accumulation of excess sphingolipids in the lysosomes of neurons, impairing neural development Autosomal Dominant Huntington’s disease Defective neural protein (huntingtin) may assemble into aggregates, causing damage to neural tissue. Hypercholesterolemia Defective LDL receptor leads to excessive cholesterol in blood and early heart attacks X-Linked Recessive Duchenne muscular Defective cytoskeletal dystrophy (DMD) protein (dystrophin) leads to impaired muscle function. Hemophilia A Defective blood clotting factor VIII leads to uncontrolled bleeding. 1/10,000 of European origin 1/122 French Canadians 1/3500 males 1–2/10,000 males In this schematic diagram, a tumor develops through repeated rounds of mutation and proliferation, giving rise eventually to a clone of fully malignant cancer cells. At each step, a single cell undergoes a mutation that either enhances cell proliferation or decreases cell death, so that its progeny become the dominant clone in the tumor. INHERITED HUMAN DISEASES • • Inherited human diseases are the phenotypic consequence of defective human genes. Although a “disease” gene may result from a new mutation that arose in the preceding generation, most cases of inherited diseases are caused by preexisting mutant alleles that have been passed from one generation to the next for many generations. The typical first step in deciphering the underlying cause of any inherited human disease is to identify the affected gene and its encoded protein. Comparison of the sequences of a disease gene and its product with those of genes and proteins whose sequence and function are known can provide clues to the molecular and cellular cause of the disease. COMMON INHERITED HUMAN DISEASES Diseases Molecular and Cellular Defect Autosomal Recessive Sickle-cell disease Abnormal hemoglobin . causes deformation of red blood cells, which can become lodged in capillaries; also confers resistance to malaria Cystic fibrosis . Defective chloride channel . (CFTR) in epithelial cells leads to excessive mucus in lungs Phenylketonuria Defective enzyme in (PKU) phenylalanine metabolism (tyrosine hydroxylase) results in excess phenylalanine leading to mental retardation, unless restricted by diet. Incidence 1/625 of subSaharan African origin 1/2500 of European origin 1/10,000 of European origin INFECTIOUS DISEASES Chlamydia trachomatis and Neisseria gonorrhoeae • These bacteria are currently the two most prevalent pathogens in the United States. The costs of infections, to individuals and society, are significant. Unfortunately, these pathogens can be difficult to detect with conventional diagnostic methods. The incidence of asymptomatic infection, its impact on individuals, and its influence on the prevalence of disease in the community has led multiple professional organizations (national and international) and federal agencies to recommend that all sexually active women aged 25 yr and younger and all other asymptomatic women at risk of infection be screened for C. trachomatis genital infection. Neisseria gonorrhoeae causes symptomatic disease and asymptomatic infection. In men, gonorrheae manifests itself within 7 day of infection as acute urethritis, with a penile discharge that ranges from scant, clear to copious and mucopurulent. HUMAN PAPILLOMAVIRUS (HPV) • Human papillomavirus is a double-stranded tumor virus that belongs to larger papovavirus family. HPV that are known to infect cutaneous or mucosal sites in humans; some viral types only infect oral, whereas others infect both oral and genital sites. Early gene proteins E6 and E7 are oncoproteins with transforming potential. These two viral proteins have been shown to bind to and inactivate cellular p53 and the Rb tumor suppressor gene products. Human papillomavirus is major cause of carcinoma and precancerous lesions of the cervix. The lesions that lead to cervical cancer are usually detectable using Pap test and are highly curable, but Pap test isn’t perfect screening test. • Forensic science is the branch of science dealing with the analysis of collected evidences during investigation of a crime scene. The branch emphasizes the manner of death and tries to find out the actual cause of death. DIFFERENCE OF FORENSIC TESTING AND CLINICAL LABORATORY TESTING • The forensic question is usually one of identify rather than one of presence or absence, character, or quantity as in most clinical laboratory analysis. Specimens receives by forensic laboratory are much more diverse than the typical blood, fluid and tissue samples handled by clinical laboratories. Clinical samples are collected under controlled circumstances, while evidence from which DNA must be isolated may be exposed to the environment in a variety of ways. Experiments may be necessary to validate testing for a particular case. Evidentiary material cannot be replenished and may be present in only trace amounts. Testing may consume the sample and thus complete or repeat testing may be impossible. The forensic identity testing is scrutinized in a judicial environment requiring complete accounting for the whereabouts of evidence following its collection and strict procedures. Biological Samples useful for Forensic testing: o Blood o Urine o Nails o Hair o Bile o gastric contents o liver o brain tissue • • HUMAN IMMUNODEFICIENCY VIRUS TYPE 1 (HIV-1) • Both HIV-1 and HIV-2 are members of the genus Lentivirinae of the family of Retroviridae. They are enveloped positive-stranded RNA viruses, and the virion contains two identical copies of the single-stranded RNA of approx 9–10 kb in length and the enzyme reverse transcriptase. The envelope contains virally encoded glycoproteins that mediate the adsorption and penetration into host cell via chemokine receptors CCR5 & CXCR4. • • HEPATITIS C VIRUS (HCV) • A number of the proteins of HCV appear to be important in HCV function or in HCV treatment. The Envelop proteins are among the most variable parts of the peptide chain and are associated with multiple molecular forms in a single infected person. These mutations seem to be critical for escape from the host immune response. The NS5a protein contains an interferon-response element; there is evidence in many studies that mutations in this gene are associated with resistance to interferon, the main antiviral agent used in the treatment of HCV. • DNA TYPING SYSTEMS AND THEIR CHARACTERISTICS CYTOMEGALOVIRUS • Cytomegalovirus (CMV) remains an important pathogen in immunocompromised individuals, including transplant recipients, persons with acquired immunodeficiency syndrome (AIDS), and those using corticosteroids or other immune-modulating drugs. Cytomegalovirus is a member of the Herpersviridae family and is a large double-stranded DNA enveloped virus, which replicates slowly and only grows in human diploid fibroblasts. Traditionally, detection and quantitation of CMV has relied on culture and antigendetection methods. LESSON 2: ROLE OF MOLECULAR BIOLOGY IN FORENSIC SCIENCE INTRODUCTION • DNA Testing has revolutionized criminalists. Only fingerprint evidence can rival the ability of DNA as trace evidence left at a scene to identify a perpetrator. As general rule, other trace evidence merely links an article, instrument, or material to a scene. The origin of DNAbased identity testing is generally traced to a 1985 article in Nature by Alec Jeffreys, DNA Fingerprint and suggested that the hybridization of DNA probes to polymorphic genetic loci could be exploited for forensic purposes. Genetic Timeframe Systems RFLP analysis 1980s-1990s PCR dot-blots 1980s-1990s STRs mtDNA Y-markers Alu repeats SNPs • PCR- Discrimination Comments based ++++ Labors intensive + + Limited discrimination Late 1990s to + ++++ Current present mainstay Mid1990s to + ++ Used in hairs, present skeletons Beginning use + ++ Male DNA Not yet in use + ++ Population marker Not yet in use + ++ Useful for samples with highly degraded DNA VNTR (Variable number of Tamdem Repeats) Analysis by RFLP (Restriction Fragment Length Polumorphisms) – most predominant method VNTR with SLP (Single Locus probe). SLP genetic systems are robust and generates readily interpretable bands, more precise allele frequency statistics, and less sensitive to fragmentation of high molecular weight DNA. • • STR (Short Tandem Repeats) - quick, less expensive and more forgiving with respect to technical skills needed and sensitive to DNA degradation. These 13 core loci (FBI Panel of Forensic Scientist) have become the standard for casework and databanking for most forensic laboratories around the world. STEPS ON HOW DNA FINGERPRINT IS PRODUCED I. DNA FINGERPRINTING 1. 2. 3. • • • DNA fingerprinting is a method used to identify an individual from a sample of DNA by looking at unique patterns in their DNA. A DNA sample taken from a crime scene is compared with a DNA sample from a suspect. If the two DNA profiles are a match, then the evidence came from that suspect. Conversely, if the two DNA profiles do not match, then the evidence cannot have come from the suspect. DNA fingerprinting is also used to establish paternity. Just like your actual fingerprint, your DNA fingerprint is something you are born with, it is unique to you. Gene Markers and Y chromosome Markers - useful as a gender marker. Males will display amelogenin locus (46 X,Y) heterozygosity and Female (46 X,X) homozygosity. Male specific DNA obtained from vaginal swab can be typed without the usual “differential extraction” in which DNA from spermatozoa is released after the female fraction from epithelial cells. Mitochondrial DNA (mtDNA) – present in hundreds to thousands of copies per cell. MtDNA is transmitted to children via oocytes. Although it is generally thought that mitochondrial DNA is exclusively derived from the mother, a minor contribution from the father is occasionally present, particularly in disease states. Primarily useful in identity testing in three contexts: 1. sample may be available that contains mitochondrial but not nuclear DNA. Example, shed hairs that do not have roots generally contain only mtDNA. 2. DNA within a specimen such as skeletal remains, is substantially degraded, the high copy number of mtDNA. 3. mtDNA analysis may become essential when only a distant relative is available for a reference specimen. The first step of DNA fingerprinting was to extract DNA from a sample of human material, usually blood. Molecular scissors called restriction enzymes were used to cut the DNA. This resulted in thousands of pieces of DNA with a variety of different lengths. These pieces of DNA were then separated according to size by a process called gel electrophoresis. o The DNA was loaded into wells at one end of a porous gel, which acted a bit like a sieve. o An electric current was applied which pulled the negatively-charged DNA through the gel. o The shorter pieces of DNA moved through the gel easiest and therefore fastest. It is more difficult for the longer pieces of DNA to move through the gel so they travelled slower. o As a result, by the time the electric current was switched off, the DNA pieces had been separated in order of size. The smallest DNA molecules were furthest away from where the original sample was loaded on to the gel. LESSON 3: DNA FINGERPRINTING INTRODUCTION • DNA fingerprinting was invented in 1984 by Professor Sir Alec Jeffreys after he realised you could detect variations in human DNA, in the form of these minisatellites. DNA fingerprinting is a laboratory technique used to establish a link between biological evidence and a suspect in a criminal investigation. Almost every cell in our body contains our DNA. 4. 5. Once the DNA had been sorted, the pieces of DNA were transferred or ‘blotted’ out of the fragile gel on to a robust piece of nylon membrane and then ‘unzipped’ to produce single strands of DNA. Next the nylon membrane was incubated with radioactive probes. Probes are small fragments of minisatellite DNA tagged with radioactive phosphorous. The probes only attach to the pieces of DNA that they are complementary to in this case they attach to the minisatellites in the genome. 6. The minisatellites that the probes have attached to were then visualised by exposing the nylon membrane to X-ray film. When exposed to radioactivity a pattern of more than 30 dark bands appeared on the film where the labelled DNA was. This pattern was the DNA fingerprint.To compare two or more different DNA fingerprints the different DNA samples were run side-by-side on the same electrophoresis gel. III. SOLVING CRIME II. DNA PROFILING • • Modern-day DNA profiling is also called STR analysis and relies on microsatellites rather than the minisatellites used in DNA fingerprinting. Microsatellites, or short tandem repeats (STRs), are the shorter relatives of minisatellites usually two to five base pairs long. Like minisatellites they are repeated many times throughout the human genome. How is a DNA profile produced • • • • • • 1. 2. 3. 4. DNA is extracted from a biological sample. STR analysis is incredibly sensitive so it only needs a tiny amount of someone’s DNA to produce an accurate result. As a result the DNA can be extracted from a wider range of biological samples including blood, saliva and hair. Unlike the original DNA fingerprinting method, DNA profiling does not use restriction enzymes to cut the DNA. Instead it uses the polymerase chain reaction (PCR) to produce many copies of specific STR sequences. 1. PCR is an automated procedure that generates lots of copies of a specific sequence of DNA. It only requires small amounts of DNA to start with and can even make copies from a DNA sample that is partially degraded. 2. In PCR small bits of DNA called primers? bind to complementary sequences of the DNA of interest and mark the starting point for the copying of the DNA of interest. 3. In STR analysis the primers used in the PCR are designed to attach to either end of the STR sequence of interest. 4. The primers for each STR is labelled with a specific coloured fluorescent tag. This makes it easier to identify and record the STR sequences after PCR. Once enough copies of the sequence have been produced by PCR, electrophoresis is used to separate the fragments according to size. Each fragment passes by a laser which causes the fragments with fluorescent tags to glow with a specific colour. The output is displayed as a series of coloured peaks highlighting the colour and length of each STR sequence. • DNA profiles are very useful in forensics because only a tiny sample of human material left behind after a crime may be sufficient to identify someone. In the UK, a complete DNA profile consists of 11 STR sequences plus a sex determiner to confirm if the profile is from a man or a woman. Now all new profiles include an additional five STR sequences to provide consistency across borders in Europe. In the USA, the Federal Bureau of Investigation (FBI) recommends that 13 STR sequences are tested. Many states are increasing the number of STR sequences tested to enable more efficient investigations across state borders. A match made between a crime scene profile and an individual profile identifies a possible suspect. A match made between different crime scene profiles indicates a repeat offender at work. The police may use this DNA evidence to support other evidence to help prosecute someone for a crime. Complete DNA profiles give very reliable matches and may provide strong evidence that a suspect is guilty or innocent of a crime. LESSON 4: GENE MAPPING INTRODUCTION • • Genetic mapping offers evidence that a disease transmitted from parent to child is linked to one or more genes and provides clues about which chromosome contains the gene and precisely where the gene lies on that chromosome. Gene mapping describes the methods used to identify the locus of a gene and the distances between genes. Genetic mapping is also called linkage mapping - can offer firm evidence that a disease transmitted from parent to child is linked to one or more genes. Mapping also provides clues about which chromosome contains the gene and precisely where the gene lies on that chromosome. Genetic maps have been used successfully to find the gene responsible for relatively rare, single-gene inherited disorders such as cystic fibrosis and Duchenne muscular dystrophy. Genetic maps are also useful in guiding scientists to the many genes that are believed to play a role in the development of more common disorders such as asthma, heart disease, diabetes, cancer, and psychiatric conditions. LESSON 5: PROBABILITY AND DNA TESTING INTRODUCTION • • In gene mapping, any sequence feature that can be faithfully distinguished from the two parents can be used as a genetic marker. Genes, in this regard, are represented by "traits" that can be faithfully distinguished between two parents. Their linkage with other genetic markers are calculated in the same way as if they are common markers and the actual gene loci are then bracketed in a region between the two nearest neighbouring markers. The entire process is then repeated by looking at more markers which target that region to map the gene neighbourhood to a higher resolution until a specific causative locus can be identified. • TYPES OF PROBABILITIES • • HOW DO RESEARCHERS CREATE A GENETIC MAP? • • Scientists isolate DNA from these samples and examine it for unique patterns that are seen only in family members who have the disease or trait. These characteristic patterns in the chemical bases that make up DNA are referred to as markers. This is why: when eggs or sperm develop, the paired chromosomes that make up a person's genome exchange stretches of DNA. Think of it as a shuffling process, called recombination. The single chromosome in a reproductive cell contains some stretches of DNA inherited from the person's mother and some from his or her father. If a particular gene is close to a DNA marker, the gene and marker will likely stay together during the recombination process, and they will likely be passed on together from parent to child. If each family member with a particular disease or trait also inherits a particular DNA marker, it is very likely that the gene responsible for the disease lies near that marker. USES OF GENE MAPPING • Identify genes responsible for disease 1. Hereditary disease 2. Cancer Each child inherits one copy of this DNA segment from the mother and one copy from the father. These DNA segments are called alleles. A DNA test result with a Probability of Paternity of 0% means that the alleged father is excluded, or cannot be the biological father. A DNA test result with a Probability of Paternity of 99% or greater means that the alleged father is most likely the biological father. Probabilities are mathematical measures of likelihood. They’re a way of quantifying (giving a specific, numerical value to) how likely something is to happen. A probability of 1 for an event means that it is guaranteed to happen, while a probability of 0 for an event means that it is guaranteed not to happen. A simple example of probability is having a ½ + ½ = 2 chance of getting heads when you flip a coin. Empirical Probability of an event is calculated by counting the number of times that event occurs and dividing it by the total number of times that event could have occurred. If the event you were looking for was a wrinkled pea seed, and you saw it 1, 850times out of the 7, 324 total seeds you examined, the empirical probability of getting a wrinkled seed would be: o Example: 1,850/7,324 = 0.253 or 1 in every 4 seeds. Theoretical Probability of an event is calculated based on information about the rules and circumstances that produce the event. It reflects the number of times an event is expectedto occur relative to the number of times it could possibly occur. o Example: if you had a pea plant heterozygous for a seed shape gene (Rr) and let it self-fertilize, you could use the rules of probability and your knowledge of genetics to predict that 1 out of every 4 offspring would get two recessive alleles (rr) and appear wrinkled, corresponding to a 0.25 (1/4) probability. RULES IN PROBABILITY THE PRODUCT RULE • • • Two events same outcome One probability rule that's very useful in genetics is the product rule, which states that the probability of two or more independent events occurring together can be calculated by multiplying the individual probabilities of the events. Example: o If you roll a six-sided dice once, you have a 1/6 chance of getting 6. o If you roll two dice at once, your chance of getting two sixes is: (probability of a six on dice 1) x (probability of a six on dice 2) = 1/6 x 1/6 = 1/36 o If both event X and event Y must happen in order for a certain outcome to occur, and if X and Y are independent of each other (don’t affect each other’s likelihood), then you can use the product rule to calculate the probability of the outcome by multiplying the probabilities of X and Y. o case 1, AF included; case 2, AF excluded; case 3, AF excluded; case 4, AF included. (Courtesy of Fairfax Identity Laboratories, a division of Commonwealth Biotechnologies, Inc.) THE SUM RULE OF PROBABILITY • • • • One event can occur more than 2 different ways) The probability that any of several mutually exclusive events will occur is equal to the sum of the events individual probabilities. Example 1: o If you roll a six-sided dice, you have a 1/6 chance of getting any given number, but you can only get one number per roll. o You could never get both a one and a six at the same time; these outcomes are mutually exclusive. o The chances of getting either a one or a six are: (probability of getting a 1) + (probability of getting a 6) 1/6 + 1/6 = 1/3. Example 2: • Electrophoresis of various biochemical markers became widely available. With this process, proteins from a person's blood or other tissue were placed onto a gel, such as potato starch, agarose, or polyacrylamide. An electric current was then run through the gel, and different forms or isozymes of the proteins were separated by their electrical charge and/or size. Differences in isozymes relate to differences in the alleles that code for these proteins GENETIC SCREENING (PPT) WHAT IS GENETIC SCREENING? • • • • o The probability of offspring with a dominant phenotype is: (probability of A from Mom and A from Dad) + (probability of A from Mom and a from Dad) + (probability of a from Mom and A from Dad) = ¼ + ¼ +¼=¾ APPLICATION: PATERNITY TESTING • • RFLP autorad results from paternity casework. DNA samples from four traditional paternity trios (M = mother, C = child, AF = alleged father) were digested with HaeIII, run on gel, and transferred to a nylon membrane via Southern blotting. The membrane was hybridized to a radioactively labeled probe (32P) for EFD52 (a VNTR variable number of tandem repeats locus). The resulting autoradiograph is shown. The first and last lanes represent molecular-weight markers. Paternity results for this locus are indicated as follows: Genetic screening is the use of simple diagnostic tests performed on a large number of individuals, to identify those who are at a high risk of having or passing on a specific genetic disorder. How does Genetic Screening differ from Genetic Testing? o The major difference between them can be explained in examining WHY an individual undergoes genetic testing If someone desires to be tested due to the possibility that he or she may have a disease gene because a large percentage of people in the same age group or ethnic group are at high risk for having the gene, this individual would need to undergo genetic screening. PURPOSES OF GENETIC SCREENING • • • • • Confirm the diagnosis in patients with unusual sign and symptoms. Detect the possible cause of the genetic/hereditary in origin For treatment and prevetion of the disease advise the family members of the diagnosed patient or carriers in the family. In the population study, screening is carried out genetic markers for the disease prevalence in groups or races MARKERS • Variations in larger pieces of DNA, which scientists search in order to locate the faulty gene. TYPES OF GENETIC SCREENING • • • • • • Carrier Screening Prenatal Screening Newborn Screening Presymptomatic Screening Forensic/identity Screening Susceptibility Screening NEWBORN SCREENING • • Identifies biochemical or other inherited conditions in newborns that may result in mental retardation or other complications. Newborn screening is an effective measure in preventing mental retardation. CARRIER SCREENING • • • It is used to describe direct gene testing applied to a whole population or to a defined group. Ex. genetic carrier screening may be available for people in the population who have no personal or family history of a condition but who have a greater than average chance of carrying a particular faulty gene due to their ancestry. Clinical and Biochemical abnormalities used in Carrier detection of X-linked disorders Disorder Abnormality Clinical Ocular Albinism Mosaic retinal pigmentary pattern Alport Syndrome Hematuria Biochemical Hemophilia A Reduced factor VIII activity antigen ratio Hemophilia B Reduced levels of factor IX G6PD deficiency Erythrocyte G6PD activity reduced Duchenne Muscular Raised serum creatinine Dystrophy kinase level PRESYMPTOMATIC SCREENING • • Used for predicting adult-onset disorders. It is also for estimating the risk of developing adult-onset genetic disorders. Ex. Huntington's disease, cancers, and Alzheimer's diseases. FORENSIC/IDENTITY SCREENING • Employed to identify suspects in criminall cases (especially murder, rape and other violent assaults) and analysis of family relationships in disputed paternity and immigration cases. PRENATAL SCREENING • Available to those individuals who have a higher risk for passing a disease gene on to the child. It is used to determine the genetic make-up of an unborn child. SUSCEPTIBILITY SCREENING • Involves the screening of selected populations for genetic susceptibility to environmental hazards. SCREENING TESTS SCREENING TESTS IN PRENATAL DIAGNOSIS • • Chorionic Villus Sampling (CVS) o Usually done at 10 to 12 weeks of pregnancy to obtain a sample of the placenta by passing a plastic tube through the vagina and into the uterus or by passing a needle through the abdomen and into the uterus. o This allows doctors to diagnose many of the same conditions as amniocentesis, but earlier in the pregnancy blood test for alpha-fetoprotein (AFP) may be performed at 16 to 18 weeks of pregnancy . o It is used to indicate the level of AFP, a substance produced by the fetus and passed to the mother's blood. • Ultrasound o usually performed as early as possible in pregnancy, and is a non-invasive procedure that provides a visual image of the fetus • Embryo biopsy o This can be used on an embryo conceived by in vitro fertilization to determine if the embryo is free of certain genetic disorders before it is implanted in the uterus. Amniocentesis o It may be done at 13 to 18 weeks of pregnancy. o It is widely-used procedure of obtaining amniotic fluid from the uterus by using a needle to pass through the abdomen. NEWBORN SCREENING TESTS Disease Phenylketonuria Congenital hypothyroidism • Maternal Serum alpha-fetoprotein (MSAFP) screening o is a blood-screening test performed at the 16-18 week gestation date and tests for Spina bifida Biotidinase deficiency Galactosemia Homocystinuria Maple syrup urine disease Sickle-cell disease Congenital adrenal hyperplasia Test/ Method Automated fluorometric assay Thyroxine or thyroid-stimulating hormone Specific enzyme assay Modified Guthrie test Modified Guthrie test Modified Guthrie test Hemoglobin electrophoresis 17- Hydroxyproesterone assay HOW IS SCREENING DONE? • • • • A blood sample is drawn and sent to a genetics laboratory for DNA analysis. Certain locations on the DNA chain are examined for mutations that are known to be linked to specific genetic conditions in defined populations. After several weeks, a report is available which shows whether or not the individual has the mutations linked to particular diseases in a given population. The report should be carefully read by a knowledgeable doctor or trained genetic counselor. A screening report is not a simple yes/no situation. It will tell the doctor or genetics specialist what the chances are that the patient is a carrier of a particular disease based on the mutations tested for. ADVANTAGES AND DISADVANTAGES OF GENETIC SCREENING ADVANTAGES • • • • Testing provides individuals with an opportunity to seek genetic counseling so that the risk of inheriting a genetic disease may be reduced. Testing can lessen the anxiety of not knowing the possibility of developing a disease and provide a tremendous sense of relief once a definitive test result is given For medical regimen, prepare for, and possibly help prevent, a genetically caused condition. For make more informed decisions in the future. DISADVANTAGES • • • Once a person has tested positive for a disease or condition, he or she risks stigmatization, loss of health or life insurance, loss of employment or educational opportunities and possibly can lose the ability to adopt a child. Privacy of other family members is threatened since, if one family member is tested, this information implies that relatives also could have the disease gene or may have an increased risk for disease. Inability to predict the severity number of unique factors. Even couples who choose prenatal testing cannot always know in advance the severity of the disability in their child. •