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by: Charles Kohler


Charles Kohler
GPA 3.94


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This 127 page Class Notes was uploaded by Charles Kohler on Tuesday October 13, 2015. The Class Notes belongs to BIOL 4190 at Louisiana State University taught by Staff in Fall. Since its upload, it has received 33 views. For similar materials see /class/222840/biol-4190-louisiana-state-university in Biological Sciences at Louisiana State University.

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Date Created: 10/13/15
CORONAVIRUS LECTURE NOTES BIOL 4190 K G KOUSOULAS Introduction Coronaviruses were first isolated from chickens in 1937 After the discovery of Rhinoviruses in the 195039s 50 of colds still could not be ascribed to known agents In 1965 Tyrrell and Bynoe used cultures of human ciliated embryonal trachea to propagate the first human coronavirus HCoV in vitro There are now approximately 15 species in this family which infect not only man but cattle pigs rodents cats dogs and birds some are serious veterinary pathogens especially chickens Group IV sense RNA Viruses Order Nidovirales quotNestedquot Viruses Family Genus Type Species Hosts Subfamily lArteriviridae Arterivirus Equine arteritis virus V Vertebrates iCoronaviridae Coronavirus infectious bronchitis virus Vertebrates l Torovirus Equine torovirus Vertebrates lRoniviridae Oka virus Gill associa ted virus Vertebrates BIOLOGICAL PROPRETIES Coronaviruses infect birds and mammals including the humans These viruses typically infect the respiratory tract and gastrointestinal organs however other organs such as the liver heart kidneys and the eyes can be affected These viruses are typically restricted to their primary host however occasionally they can infect other animal species ie bovine coronavirus infection of turkeys and humans and experimental infection of dogs by transmissivble gastroenteritis virus TGEV The most famous example of this crossing of hostbarrier is the SARS Respiratory Syndrome Acute Syndrome virus which was potentially transmitted from the civets to humans Morphology Coronavirus particles are irregularly shaped 60 220nm in diameter with an outer envelope bearing distinctive 39club shaped39 peplomers 20nm long x 10nm at wide distal end This 39crown like39 appearance Latin corona gives the family its name The centre of the particle appears amorphous in negatively stained EM preps the nucleocapsid being in a loosely wound rather disordered state The envelope carries three glycoproteins S Spike protein receptor binding cell fusion major antigen E Envelope protein small envelope associated protein M Membrane protein transmembrane budding amp envelope formation induction of interferon responses In a few virus types there is a third glycoprotein HE Haemagglutinin esterase The genome is associated with a basic phosphoprotein N N modulates viral RNA synthesis binds to viral RNA and forms the genomic helical structure Relative number of structural in each Virion Protein Coramzvirus Taravims 180220 200 Cellular receptors The coronavirus are divided in 3 groups The members of the group 1 of the coronavirus include TGEV HCoV229E use aminopeptidase N APN or CD13 as a receptor for entering into cells Group 2 coronaviruses MHV use the carcionoembryonic antigen CEA as a receptor Also BCoV and HCVOC43 may use N acetyl90acetylneuraminic acid moieties for binding to cell surfaces The SARC CoV uses the angotensin2 ACE2 as a receptor The coronavirus virion swim glympml m Hemagglwmmyleslmse V glympmlem Membrane glymprmem mu mi gimme Nucleanps plmspmwm Genome Nonsegmented singlestranded sense RNA 2731 kb dependent on virus the longest ofany RNA virus The genome has a 539 methylated cap and 339 polyA and functions directly as mRNA Following the 539 methylated cap there is a leader sequence of65 to 98 nucl otides and a nontranslated region of 200 to 400 nucleotides In the 339 terminus of the viral genome there is also a non translated sequence of 200 to 500 nucleotides followed by a tail The virion RNA contains 7 to 10 functional genes of which 4 or 5 encode for the structural proteins The genes are arranged in a precise order 539Polym raseHE SEMN3 39 with a variable number of other genes that are apparently nonstructural ns and often non essential for replication of the virus in tissue culture About 23 of the viral RNA is occupied by the polymerase gene The polymerase gene includes two open reading frames that in some cases are overlapping or very proximal to each other There is a quotslipperyquot slipping nucleotide sequence of 7 nucleotides and a pseudoknot structure which serves as a signal for a translational shift of the ribosomal machinery to read the second open reading frame 18 Replication Q receptor Genomic RNA 39 Negative Strand Template 39 V smoothwalled RNA Proteins T les ti 2 Ns2 V98 quotquotN2 139gtHE39 3 5 39 pJ6M o7 N 3 l gt Pol F o nucleocopsid 65 4 NS4 X Golgi mRNA S h N 5 E Genomic RNA Initially the 539 ZOkb of the sense genome is translated to produce a viral polymerase which then produces a fulllength sense strand this step is poorly understood This is used as a template to produce mRNA as a 39nested set39 of transcripts all with an identical 539 nontranslated leader sequence of 72nt amp coincident 339 polyadenylated ends Initially the 539 ZOkb of the sense genome is translated to produce a viral polymerase which then produces a fulllength sense strand this step is poorly understood This is used as a template to produce mRNA as a 39nested set39 of transcripts all with an identical 539 nontranslated leader sequence of 7Znt amp coincident 339 polyadenylated ends Genome RNA 32 kb gene 1 replicase genes 27 V i i z H v Leader replicase polyprolein WV Each mRNA is monocistronic the genes at the 539 end being translated from the longest mRNA amp so on These unusual cytoplasmic structures are produced not by splicing posttranscriptional modification but by the polymerase during transcription Between each of the genes there is a repeated intergenic sequence UCUAAAC which interacts with the transcriptase plus cellular factors to 39splice39 the leader sequence onto the start of each ORF Assembly occurs by budding into the golgi apparatus particles being transported to the surface of the cell by the secretory nature of this organelle amp released Pathogenesis Coronaviruses infect a variety of mammals amp birds The exact number of human isolates are not known as many cannot be grown in culture In humans they Respiratory infections common including Severe Acute Respiratory Syndrome SARS Enteric infections occasional mostly in infants lt12 months Neurological syndromes rare They are transmitted by aerosols of respiratory secretions by the fecaloral route and by mechanical transmission Most virus growth occurs in epithelial cells Occasionally the liver kidneys heart or eyes may be infected as well as other cell types such as macrophages In coldtype respiratory infections growth appears to be localized to the epithelium of the upper respiratory tract but there is no adequate animal model for the human respiratory coronaviruses Clinically most infections cause a mild selflimited disease classical 39cold39 or upset stomach but there may be rare neurological complications SARS is a form of viral pneumonia where infection encompasses the lower respiratory tract Coronavirus infection is very common and occurs worldwide The incidence of infection is strongly seasonal with the greatest incidence in children in winter Adult infections are less common The number of coronavirus serotypes and the extent of antigenic variation is unknown Reinfections appear to occur throughout life implying multiple serotypes at least four are known andor antigenic variation hence the prospects for immunization appear bleak 4 quot7 D g SARS is a type of viral pneumonia with symptoms including fever a dry cough dyspnea shortness of breath headache and hypoxaemia low blood oxygen concentration Typical laboratory findings include lymphopaenia reduced lymphocyte numbers and mildly elevated aminotransferase levels indicating liver damage Death may result from progressive respiratory failure due to alveolar damage The typical clinical course of SARS involves an improvement in symptoms during the first week of infection followed by a worsening during the second week Studies indicate that this worsening may be related to patient39s immune responses rather than uncontrolled viral replication The outbreak is believed to have originated in February 2003 in the Guangdong province of China where 300 people became ill and at least five died After initial reports that a paramyxovirus was responsible the true cause appears to be a novel coronavirus with some unusual properties For one thing the SARS virus can be grown in Vero cells a fibroblast cell line isolated in 1962 from a primate a novel property for HCoV39s most of which cannot be cultivated In these cells virus infection results in a cytopathic effect and budding of coronaviruslike particles from the endoplasmic reticulum within infected cells The SARS virus is believed to be spread by droplets produced by coughing and sneezing but other routes of infection may also be involved such as faecal contamination The most common reported symptom is fever 94 with 51 72 of patients reporting general influenzalike symptoms chills malaise loss of appetite and myalgia Gastrointestinal symptoms are less common at presentation including diarrhoea 27 vomiting 14 and abdominal pain 13 The mean incubation period otSARS is estirnated to be 6 4 davs The estirnated case tatahtv rate is 13 2 tor patients vounger than 60 vears and 43 3 tor patients aged 60 vears or oider Amphtication of short regions ot the powrnerase gene mum the rnost strongw conserved part of the coronawrus rn v rever e transcriptase powrnerase chain reaction memo and nucieotide sequencing reveaied mums that the SARS virus is a nove coronavirus which has wequot not prewousw been present in rnan i i V rn GENPHI coronaviruses with no obwousw unusuai teatures msm o gh there are sorne di e e rn ot the monastructurd proteins which are unusuai E 339 c 5mmquot Hawnull There is currentw no generai agreernent that antwirai drugs have been shown to be consistentw successtui in treating SARS or anv coronawrus intection Diagnostic tests tor coronawrus intection taii into two tvpes Serological testing for antiacoronavirus antibodies consists of indirect H r tin A r r A i i A h t ELISA which detect antibodies against the wrus produced in response to infection Aithough so patients have detectabie coronawrus antibodv within 14 davs ot iiiness onset detinitwe interpretation otnegatwe coronavirus antibodv tests is possibie onw tor specirnens obtained gt21 davs after onset of fever 3 0 Molecular testing consists of reverse transcriptaseapoivmerase chain reaction RTVDCR tests speci c for the RNA trorn this novei coronavirus This a etect intection within the first 10 davs after the onset ottever in sorne SARS patients but the duration otdetectabie wraernia and VH US shedding is unknown so RTVDCR tests pertorrned too iate couid gwe negatwe resuits Cornrnerciai diagnostic tests are now avaiabe Origin of the SARS virus Coronaviruses with 99 sequence sirniiarity to the surface Spike protein othurnan SARS isoiates have been isoiated in Guangdong china trorn apparentw heaithv masked pairn cwets Paguma larvata a catahke rnarnrnai ciosew reiated to the rnongoose The uniucw pairn cwet is regarded as a dehcacv in Guangdong and it is believed that humans became infected as they raised and slaughtered the animals rather than by consumption of infected meat Might SARS coronavirus recombine with other human coronaviruses to produce an even more deadly virus Fortunately the coronaviruses of which we are aware indicate that recombination has not occurred between viruses of different groups only within a group so recombination does not seem likely given the distance between the SARS virus and HCoV There is considerable experience of development of coronavirus vaccines for veterinary purposes though not all of it is encouraging On the whole inactivated coronavirus vaccines induce poor protection The spike protein alone can induce immunity but the internal nucleoprotein has also been reported to induce protective immunity The WHO has recommended that SARS vaccines be developed The quickest and probably safest to develop would be an inactivated or subunit vaccine Even if such a vaccine were not fully protective against SARS infection it might still provide some protection against lifethreatening SARS pneumonia Gene Therapy Strategy fortransfer of a gene to a patient cDNA protein coding sequence Plasmid or Regulatory reion promoter and enhancer vim vector determines tissuespeci city determines amount of expression allows cDNA to be regulated Transfer DNA directly Transfer DNA into into patient patient39s cells in culture Patient Three categories of somatic cell gene therapy I Ex vivo 7 cells removed from body incubated with vector and geneengineered cells returned to body 2 In situ 7 vector is placed directly into the affected tissues 3 In vivo 7 vector injected directly into the blood stream Indirect Ex Vivo DirecT In Vivo Viral Q g rol 1 Au gar Targeted Cell J K NonViral NonVlrol Example of ex vivo somatic cell gene therapy 0 Usually done With blood cells because they are easiest to remove and return Sickle cell anemia Example of in situ somatic cell gene therapy Infusion of adenoviral vectors into the trachea and bronchi of cystic fibrosis patients 0 Injection of a tumor mass with a vector carrying the gene for a cytokine or toxin 0 Injection of a dystrophin gene directly into the muscle of muscular dystrophy patients Example of mvz vo somatic cell gene therapy 0 No clinical examples 0 In vivo inj ectable vectors must be developed Conecu39ve gene construct integration Into gen Expansion ofengineered cells and implantation into patient extraceHu ar mteracuons 59 i a 2 E don and maintenance 0 cient msfecu39on and transp I om exogenous DNA gene transfer vector plasma membrane ftarget cells on ofDNA to nucleus for mnsgene nuc eus gt expressmn cymsm g nuc eases degradation gt endosome gt lysnsome Types of vectors RNA viruses etrovirus 1 Murineleukemn 2 39 es ms mmunode cienc DNA virus 1 Adsnovim 2 Adenoas 3 Herpes simplex m H 4 Pox viruses vi Human y mes luv 3 Human Tcelllymphou39opicviruses HTLV es 5 sociated viruses AAV 39 39 s sv 3 Liposomepolyca on complexes 4 Peptide dcliverysystems Life cycle of a retrovirus inglurstmndud mm mm plugmw m punkm Mhzunmmlng man quotW39M mummm amid mm mm r t m W a m onva mm mm mm tssmnu cuLnNnIoss mum r 4W1 r i V 4 5 r M 5 Lussor vmmsm mm um P YRA Nswnou J mnsmm rmnscmmmuswow m csn Rmmwmuss MAKESMANYRNACUPVES meanmm or nu momma DNA Wham cowwtowosr constructs mmWWW w Numbmquot vavNADousLtniuxsv maintained at mm m scnwus m Wm thls stage quotWquot mmm a f Vira panicls b v F I I d m n m rcvlna asmi r Ir I u p 1 j Tlanslem un Q Wecuun Reverse transcrlphcn lntegratmn Integrated pvcvirus Infected cell Transcripliunsphcing Translatmn 0f genum c and subgonomvc RNA mm 7 gag pm and env me n of viral pamcles Renewal vctar r repllcatlnn ncompetenl Repllcanuncompmem elmvlvus lnfecuun um nu replication gt m Advantages Diagram of a Betrovirus l Randomly integrates into genome 2 Wide host range 3 Long term expression of transgene Disadvantages 1 Capacity to carry therapeutic genes is small 2 lnfectivity limited to dividing cells 3 lnactivated by complement cascade 4 Safety A problem with retroviral vectors they insert randomly into the genome Using a murine leukemia virus vector the y C constant chain was inserted into 10 boys with SCID no B or T cells These are the bubble boys Removed bone marrow cells from the boys purified CD34 cells from that population transduced cells ex vivo and reintroduced cel s Startling success all boys now had an adaptive immune system Allwent home and started living more or less normal lives Science 0 Summer 2001 two youngest boys that were given greatest numbers of virions came down with an acute T cell leukemia Both were successfully treated for the leukemia Analysis of the tumor cell a clonal population of pre T cells that had the viral vector near aknown T cell oncogene thathappens to be a specific pre T cell transcription factor LMOS Smoking gun evidence that retroviral vector in the two boys had inserted into a location that stimulated expression of LMOS causing a selective advantage to those T cells and induction ofa leukemia Adenovirus Adenovirus 4 Fi bre kCapsid kknob Advantages l Ef ciency of transduction is high 2 High level gene expression 3 Slightly increased capacity for exogenous DNA Disadvantages 1 Expression may be transient 2 Cellspecific targeting dif cult to achieve 3 Virus uptake is ubiquitous 4 Safety Adenoviruses as delivery systems most use adenovirus 5 infects dividing and non dividing cells can enter many different cell types because it uses arelatively ubiquitous receptor for entry CAR produces very high titers of viml particles Nonenveloped virus 7 relatively stable virion Gnnamn smears ELquot139Jr 9quot n enovims vectors the essennal genes El andmm have been deleted andthe n at replication competent viruses as well aswildtypevimses TP acts as a primer for initiation of synthesis DBP e a DNAebinding protein DNA Pol e 140kD DNAe dependent polymerase 5 ac mm 1 Ants e we fur new 5mm ac 39dc c 3 1 S Nrwslmml c i a as 0 AC 5 339 39The viral genome is coated with DBP 39DBP helps binding of NFL INFIII also binds at a speci c recognition site between nucleotides 39 and 48 39Proteineprotein interactions between NFI and pol and pTP and NFIII help recruit the pTPepol Creation of a defective n nsduclble vmon quotmid w 1 containing the gene f 39 st m ere Tnnsreemn af ncambxmm an e hgh mar recumbmzntvn39us 5 y Innml x ufrecummznt w W n w vxmsestucel hansdcegt9U a noune i 4q m m M01mu1npneny ufmfecnun number ufmfemluus urtxansducable e m mgquot m m e Thzse cells camam Adamml El and E3 1112mm wnhm hz c2 hm between 1 and mu MOI mu m mumum Hg 5 A mm mm mm mummy gm of m suns m m a m Wus 2m Wm game mm In ted m a m n the mm mm m yum A m namzls nnz mm x m gem Drawbacks of using adenoviruses transient expression adenovirus is not stably maintained in cells 7 expression is usually lost over a several week period adenoviruses are highly immunogenic cannot effectively introduce multiple rounds of the same adenovirus serotype in vivo but there are several different serotypes of adenoviruses Would need to put your gene of interest into each different serotype if you wanted to perform multiple transductions in viva can be pathogenic 71999 University of Pennsylvania gene therapy death using the largest quantity of Ad5 vector into a human 7 severe immune response and coagulation with no evidence of gene expression productive viral replication is highly cytolytic to the infected cell in vivo delivery system use defective virus Other viral vectors 0 Adenoassociated virus infects Wide range of cells both dividing and nondividing able to integrate into host genome not associated with any human disease high efficiency of transduction Herpes simplex virus vaccinia virus syndbis virus Onyx virus limited replicating adenovirus that replicates mainly in tumor cells PARVOVIRIDAE BIOLOGY 1 Small nonenveloped icosahedral viruses with NS kb ssDNA genome some infect warmblooded animals others infect insects 2 Parvoviruses are important pathogens of dogs and cats where they destroy dividing cells of the immune system parvoviruses reguire actively dividing cells for replication no host stimulation proteins 3 Adenoassociated virus AAV is a well known defective parvovirus dependovirus subgroup that relies on coinfection with adenovirus or herpes virus as helper to stimulate host cell division 4 AAV has potential as a human therapeutic vector because its genome integrates at only one speci c site in the human genome chromosome 19 infection by itself integration frequency gt 70 Receplur mmmg amp elvqu Oj ruthroucgle Pnnlvqen tramucaunn at DNA to quotmm Ewdwu to mm DNA PATHDEENESIS7 NS Prulem nxnresainn 39 5 CAP pmtem expresav on 0 4 O Eennme renltcutmn Cupsquot maeth Dell M551 5 was Parvovirus replication RQlication occurs in the nucleus amp is thought to follow the scheme to the e All Paxvovimses are highlx replication to occur The defective viruses are helper virus for replication PARVOVIRUS REPLICATION 1 Only two ORF s rep and capsid protein translated from three families of 3 coterminal Admm mamm spliced transcripts Rm Rama Hananme 2 Replication uses host DNA Oil DO polymerase but mechanism is D m 40 ED m m unigue priming relies on haippin b l elmn l l l lwl l l J lo01 formed by terminal E inverted repeat seguences 120 5 330 bases WH 3 Rep protein is required for DNA IsymlleSiS makes sped e Rep78 amp Rep68 both have DNA binding n1cks 1n halppln loops 139 Rep IS endonuclease amp helicase activity also reguired for integpation Source Dr Perrault s lecture notes Adenovirus associated Virus AAV Advantages not associated with disease can obtain high titered virus stocks 109 lOlOml small genome that is easy to manipulate stable integration into the genome in chromosome 19 infects both dividing and nondividing cells Disadvantages can only contain 45 kb of DNA Adenovilus associatedltygti1us life cycle o 39 eren serotypes of virus that have differing cellular tropism r rir innmun a viv lI I M mm Contains 2 genes that generate a Vle total of7proteins 4 non structural mmg notinvolvedin vinon structure quot quotquot m 39 and3 structural involved in virion tructure Use of AAV as a gene delivery agent Emu MA Required adenovirus proteins E1 m Eza E4 VA Ratherthan using Ad W W helper plasrnid can use adenovuus infected cells but can have dif culties with Ad contarninated AAV plasmid mv helper plasmid Ad helper nllmld stocks Packaged FKZIIDfUCLlunl vector contains no V Highly tmnsfectable 293 genes just cells already contain the t R adenovirus gene E1 lloslosll Q rMV Can generate Very high titered stocks Ability to use multiple AAV constructs to generate proteins that are encoded by genes larger than 45 kb Package half of geneofinterest in one particle and the other half in a different particle and c0 transduce cells Approach is relatively inefficient but does work Via transplicing of RNA that is generated HERPES SIMPLEX VECTORS Viral DNA Figure 1 Flaw mm depicting me 75de nucleus loss m E and 1 gene pmriucls nnd ramquot m pmduce infectious Vimsi 5mm cm and mm mmadima early expressing cell lines my cnmplemam liege gene pmdncrs a a a rassmn m m mus Full demls xvi he cnsnucucn nr replmrmmr sential Nonessential dz nem vimsts may be mum in viaa in vim m the text and references CPA lch I ICP47 V Immuna evasion Early gana Viral DNAfpncanun HERPES SIMPLEX GENOME I b UL b HE 5 Us c amp HSV l lt lt ICPU U124 U111 gt lt ICP ICI 4 CPU ICP47 ICP4 L U mL quot1 usma HSVl icnnnw IR 2 ucnm uus L w LAT 2 mm ys 1 5 new m LS Kb W I J xx 39 I 1 TREATMENT OF CARCINOMA WITH HSV Plus ccv mmmmm mjccnnn m m nf nanm mm 2 Endmmm nfwnlnncnl mmnmlmls mm aswk mlnn mum cells mmy mmm In sume am vmmnm wrunme xi ay 2 m m m wum m may mawnm Wm mud will mass hul39reralun m m mm mm m c max mm mm x 5 myquot lawnng The mm mm mwnume Illuminati 17mm uqu Pnnum E am 1 mm 11mm 11 5mm 1 Emmy v m MenLIKE m um 1st mm mm gam mm rm cnlnmchl mnmnmamua Duncan cmnnmmm Gum m mum mam mm In hm permnnnn r me mm Mum sums Umvemly 11622 Edam m1 nrswdmm mu Poxvirus life cycle early middle and late gene expression lmd W c to lasm DNA mquot y p 6 nlsmwd quotIRMA Ilzchmenl nnlry manna lawman a a gt 3 gt Inlnmnscny an V 2 I s mums 5 we 7 J WNW My RNApolymtnsu 9e wdwm9 I amm Nl vanscnmmnlanov H mm a mom enzyme eany mRNA My WW gamma 9 are enzymes army mansmpnnn lemurs slmnlmal Jrmews mmlzma resdulmn DNA packagmg E ussumW lt7 I39m lnhmmnwucnhuctlnxnlmwlmm m xmmmmmm L A puri ed naked viral DNA is not infectious Poxviruses family members smallpox cowpox vaccinia monkey pox canary pox large highly complex and understudied viruses only family of DNA viruses that replicates in cytoplasm of cell vaccinia is used as vaccine against smallpox highly immunogenic vaccinia has very very very broad tropism for different cells from different species through homologous recombination in vaccinia infected cells can introduce genes into the virus allowing production of large quantities of recombinant protein from infected cells serve as delivery system for recombinant vaccines Muss1996FNAS93 11341 Homologous recombination in the vaccinia Virus system Plasmid containing Vaccinia promoter several available depending on when you want your gene of interest expressed in the infected cells 7 early intermediate or late in infection and a portion of the vaccinia genome that will be targeted for insertion clone in gene of interest behind the promoter up to 25 kb plasmid frequently contains some sort of selectable marker eukaryotic antibiotic Bgalactosidase etc transfect into vaccinia expressing cells and homologous recombination Will occur about 01 of the time POXVIRUS VECTORS mwm mm m mammal Tnnstulmn Mum mum mamhlulmn in mm quotus a math m NonViral vectors 1 Liposome 2 Cationic polymers W 39 397 a Wvawvv39eu v KY quot l J 3 Naked DNA 4 Peptidemediated gene delivery May overcome limitations with ViIuses including small capacity for therapeutic DNA dif culty in celltype targeting and safety conceIns my mndilinns mum Mm N5 Emu my vm Esauq 1c aneume m sun 451 a min 3ranhs av mm mm dam aquarig Lgmd HSVrLk Admuvmu 2 mamas m ishuryla mm vm mt Ed 1 mu Hunganumdasu Naawhcplut inn Simdams Ymgmqs Reinvent 1mm Pmskzlc pm Slunn 1 1 am mam m r hand a n CMrijILJ 3 smnd Em C wwhmm maa cum pm yap rm 7 cm Slumxuxiabn mp1 M c 1 mm m st Pu m HErnHt a 1 1997 Ammm A L um mm a 1 may a mug R new mm m was Hm u 1 mm mm 29 mnmmphx arm me n ma mumm ya 3 new mum m nmensc mun veclnr pm A c m mm mmrxlld mmy u nmmrm m vuuml M n funan mm any mm m m nanwnul mm mm Chm us mm mmg a 5mm M1hmnu Andaman W 7 gm have mile Iumm mm mm haw mm mm s mum an Lhe m tnnsrernfan mums mun m cn39rquucl Gene 1m a mum mm mm m pcmumnll mm the 21 Examples of Gene Therapy Trials 0 Adenosine deaminase gene transfer to treat Severe Combined lmmunoDeficiency SCID CFTR gene transfer to treat Cystic Fibrosis CF Advanced Central Nervous System CNS Malignancy Mesothelioma Ornithine Transcarbamylase Deficiency Hemophilia Sickle CellDisease Targeting of different organs by viral vectors Adamqu AN Tumors nemsmpmeuc t We muscle cells reunal k I rumors Renoqu UDDSON umors mm ennapsulalea hemawpmelxe alphavlms x aeHvery Lemmms CNS lwet mums rpes smlplsx vlms cns PNSmusnle hemampmeac are cells mews m Bralecnrmlcgy 22 One developing technology that may be utilized for gene therapy is nuclear transfer cloning El Emclealed Repmgram ming CE K a Nuclear Tram fer a Sumati Sell Harvey II Paliant II Labnratmy Process to Derive Histncompalibla Human Pluriputent Slam Calls Himocompatible Human Pluripo39nenl Elem Calls a Extend Replieatiiva Lifespan with Telemerasa G Di erenliate lnm Desired Gents 23 Ethical Considerations Use of technology for nondisease conditions such as functional enhancement or cosmetic purposes for example treatment of baldness by gene transfer into follicle cells larger size from growth hormone gene increased muscle mass from dystrophin gene In utero somatic gene therapy only serious disease should be targeted and riskbenefit ratios for mother and fetus should be favorable Potential for real abuse exists by combining cloning and genetic engineering Assigned Paper Review 24 Tame Cums ma usmg oanm Wus Name Geneva Shawnas Dmease pm ovims LAdv ElBSSkDa de e on Head amp neckcancer mu Ovaman cancer War mmm5 Hi Pancreaer cancer HI 5207 LHsvrn LacZ mmm mo mp5 gene demon m now comes or 73 Mahgnsmghoma rru Nvmzo Hsvmz 70mm m mum Hm mm acre 013 mm regmn mm mm aw Memsxahc hver mmor exogenous copy m m gene under mm m HSVrle pmmmer and a 52kt vsgmem or HSVVE DNA cncovEx mus sums msemon mm mm and mm m mm mm m was 5 Breast cancer LN HSVVH Head L neck cancer H Me ananla LN m6 HSV4 Delanoquot or new cums m ms Mahgnam gum Hm LHSVVU Namav mulanun or me and m uL and mum re5uumg m 1055 av ULSB Brass cancer 1 cvms MW Reguvauon av EIA under the PSA Promoter E3 de enan Proslale cancer HY cvm Adv Regma on mam underme rm prubssm promder and as underme human PSA Pro5taze cancer Hi ummoten wumpe E3 VanmmarGMrCSF lnsemon av EMVESF and Lscz gene5 mm W51 TK ucu5 Me syams m wm NDV Natura y gamma Advanned 50m calmer Mechanisms of nnruly c e ecl in tumor cell mum gnhun39ng at 391 cell mquot 39 quot quot quotquot E vmccmive mphcmnn In neighbor cell unruuim one MINquot 3 13W 1 i it n if a i 39 O I 4r A l L 4 umppmmutvims ml dhlunchwn mm by muulirm ol39vmu amt hhviking hy apmumw mth nfnm mal cell lwvvmhpamclv 0 1mmva mm Fyxmmucpmkln 4 r 2 Dmuqmoximymm b3v1ralprucins 39I39mnsgumcxpmxsiun Lamas mam cumbinalmn Andangiugcncsh mg endusa ll swam a cylmmic 39r lyulphncyki 25 Uldllun of 1n 1 n uphuu nlmn p l P rolein e x39 chZa gt SIMoff of synthesis 39 pmrem synthesis Flgura 3 can praialn kinaya wept PM is sl39mulated by HSV1 mamm and summed Dy autophospho hon Adivaiad PKR phasphmylalus elFZx Plusmurylabd HIP2a sum on inta mu HSV 1 3944 pmsphat Jsah mpmsmarylaaes EIFQQ and anaws grulein 5m has Gum 7mm 7mm 0mm ms mumv cammmmamm m pamam min and 35M mm cEAm Va Mm Panama and mm mm WNW m mm mm mm 5 73 Ease Pmcmatc W4 and gm cm was mm Ascmav we savanna mun pmm m m4 caravans mum Alwmmvmmcmg 73mm 3va ma Piude mam E Zym Mmmmam 7 Manama WWWan A Enumch mubuntgucmmug 7 Emmc cd quotWM uvmymm 32 Emowmmmm 7 5 F wmumw 11mm mamas MM mama 7 E wusm w hasImam mammem 7 mm m mummmmm Cymcwum pm zal 5 Hmmu w 7 mmmn Wu ame Gemmam 7 Gamayquot mmwm Tinmama Mum 26 Good reviews on the topics Baker 2004 Progress in Biophys Mol Biol 84 279 7 good review on in vivo delivery systems 7 both nonviral and some viral Ad and AAV only Focus is on cardiology 1 2 Lundstrom 2003 Trends in Biotechnology 21117 Good general review 9 Sanders 2002 Current Opinions in Biotechnology 13 437 Good review on advantages and techniques of pseudotyping retroviruses 4 Thomas et al 2003 Nature Reviews Genetics 4 346 Really nice overview 27 Genetics of Cancer Viral Oncogenesis Regulation of cell division in normal cells Cellular Transformation When the cell fails to respond to such signals it is out of control and can continue to divide The daughter cells will also continue to divide Ultimately might result in cancer Malignant Tumors Invasive and Metastasis Microscopic Examination mitotic activity high nucleuscytoplasm rati rominent nucleoli little specialized structure cells grow as a benign tumor in epithelium breakthrough basal lamina invade capillary Prlmary T m 9 4 a travel through bloodstream m n39 w less than 39I in 1000 cells 39 will survive to form metastasesl b 39 I 0 v 5 7 v 999 i adhere to blood vessel escape from blood vessel proliferate to form wall in liver Ilextravasationl metastasis in liver Clonal Carcinoma Skin Cancer Basal Skin Carcinoma Sarcoma in bone muscle or other soft tissue MEDtAL LESIONS LATERAL LESIONS 5 mm rswa r x s Leukemia Cancer of the bone marrow Too many Ieucocytes are produced Lymphoma cancer of the lymph tissue Myeloma cancer of the myeloid tissue Myeloid tissue is where blood forms in the red bone marrow in vertebrates It is important in the immune system Oncogenes A gene that is capable of transforming a normal cell into a cancerous cell Oncogenes result from the mutation of normal genes protooncogenes Oncogenes are also seen In oncogenic Viruses Viral oncogenes are derived from normal host genes that have become incorporated into the viral genome and subsequently undergo mutation Genes and Cancer Mutations that result in cancer typically occur in 3 types of genes Protooncogenes genes Whose products stimulate cell multiplication Tumorsuppressor genes genes whose products inhibit cell multiplication M utator genes genes Whose products ensure accurate DNA replication and DNA repair Regulation of Cellular Prolifieration Directly suppress cell proliferation Tumor RB p53 APC NFI Suppressor Maintain de integrity of the genome BRCA12 Genes and Cancer Retinoblastoma model for retinoblastoma Knudson s 2hit mutation a swam b Heredimry velinablnuoma mlinoblaslama Relinucell l quot RFRD39 x ll m ax bin I UVKL norma HIRH lnherlled can growth RH muiatian Firs normal cell gmwm mmn un l on y m Secoqd mnlatinns quot39quotm39quot REIRH inomIII ell growth Second mnlallon mam xa lam i i J RHRH Ioss ul mum loss nl growlh cantml growih conval l Eye tumor Eye Inmor 15 i 52 DNYSFON 7 M PHASE rm L031 3 24 mm 5 39 nuclear 39 division cgtokinesis BZPHASE cgmpiasmic Gunman 39 39 5 WTERPHASE NTERFHASE I31 PHASE S PHASE DNA replication 1998 GHRLRND PUBLISHING The cell division cycle Is all DNA I EDllCalg i is cell Dig snuugh7 B2 CHECKPD l NT ENTER There are checkpoints during Which the cell checks Whether to continue progressing through the cell cycle m tHEEKPDlNT is call mg enuugm is anvir nmanl lavarauls is DNA damaged mm ERMRND PWIlSNINE Multiple cdks and cyclins regulate passage of mammalian cells through the cell cycle Cdk lrcvclin B Cdk1cvclin A CdkAcvc in Cdkscvc l a mu Resmcnun point Restriction point Is Cdkzrcyclin A analogous to START Cdecyclin E lNITlATES mIIDUE Edk PH d W m m 7 mnuu 3932 D 51 S phase E k S J M S phaae gclm lNlTIATES 1993 gamma PususmNu There are Glspecific and G2speci c cyclins and several Cdks one to stimulate the initiation of Sphase and one for Mphase 19 M PHASE my mum whan w amp 3 mm waded I I I Wm mum Wm Cyclin degradation is part of the cellcycle regulation 52 Notice the balance between gin 5 Wm m synthes1s degradation in this g 31 waded 5 We Wm 0 Ch QQ w 5 phase cychn D 5 We cg fnmk WNW my DNA repncaimn mmm 20 5 PHASE inactivE ps3 AETiVATiDN r DNAw F i 953 m At WE p53 arms In DFpI GENE u r 2 12 2 TRANSERiPTiDN D21 mRNA wmwuw DNA damage v D2i uk mmmtur protein es9L AETWE Am 5 We WWW pm We WWW mm mm a EEEEEEEEEEEEEEEEE in p53 is a tumor suppressor brakes It regulates transcription of a Cdk inhibitor p21 21 Passage through the restriction point depends on activation of E2F transcription factors Mid lt31 Late G1 Mammalian cyclinkinase inhibitors also contribute to cell cycle control gruwth Yactur mach gmwm acuvated gruwth mum receptur Yactur receutur wtraceuumr mm 539 2 gnahng WWW actwated 5 Phase cychnrtdk mactwated cumpvex RD prutem actwe RD Drutem actwe RD Drutem mactwated gene BEWE gene reguvatury prutem reawaturu urutem A37 k mactwated gene regmamrg prutem g PHuspHuRvLATmN TRANSDR PT UN D RD i TRANSLAT DN SHEEP Z PRUUFERAUDN A quot E sthEQEEVLL aw uwgmrwq k em wkan vuausuwn How RB a tumor supressor works with Cdk 23 The p53 Signaling Pathway Mg mummy mm mm 0 Apupm p53 Dkgvadalmn i39i Kquot CeHCycle 24 Effects of DNA damage and normal nunmutant D53 lead to cell growth arrest gtgxcltion 0 V i i U Stabilization ol 53 by un u u 4 m Promoter Cy by Cdk Cdk Kinuse activiiy No e W Absence ol kinase a Iy blacks on to 3 Gt I 5 25 Arrest in a Rule of pRB in regulating cell diVIsmn a Manual cell b Cell with two mutant c Cell infected with certain DNA RE alleles tumor viruses T unshhlo pRE i cannot blnd on net runmtau and a Viral pmteins tainutoplie and r 39 52F Pt Liar 52 on ctivutos genus cuntmllau by E2F Phosphnrylallon ol pRH by cycllnlcak cuntrollad by E2F n h I mRNAs transcribed mHNAs lmnscrlbed DF1 p gsgam i d tram activated genes tram activated genes Frutaill products mad Pmleln pmducu made that am moan for that are needed lor ntry into 5 entry inlo s Viralinduced 6 5 call division 8 Released E2FDP1 acllvales genes controlled by EZF mam nsurlbed tram acllvated genes Protein products mace trial are needed lor entry n a rat s AETWATE murme AETWATE mumn 5 phase 5 phase mytuuc mvtuuc cychnrtdk cychnr dk cychnr dk cychnr dk T cumpvms cumpvms T T cumpvmsT cumpvms n da 39 u t a 2 wcumm tew cngga wanealga 5122 DNA unvavur nve Extracevm ananvhunmgm em wkan WSW 27 Tumor eruses For most viruses Genome gt viral proteins Replication gt Lysis gt Progeny virions Lytic Life Cycle 28 TABLE IJ ZViruses Associated with Human Cancers VIRUS ASSOCIATED TUMORS AREAS OF HIGH INClDENCE DNA Wrusex Papovavirus family Pap39 omavirus wans benign many distinct carcinoma of the uterine worldwide su ains ceivix Hepadnavirus family Hepatitis B virus Liver cancer hepatocellular carcinoma Herpesvirus family EpsteimBan vims Burkin s lymphoma cancer of B lymphocytes nasupharyngcal carcinoma RNA Llll39llSES Reu m lrusfai 39ly Human Trcell adult Trcell leukemia leukamiavirus lymphoma WpellHTLVJ Human immunoA Kaposi39s sarcoma de ciency Virus HIV the AIDS virus SumheaslAsia tropical Africa West Africa Papua New nine southern Ch ina Greenland lapuuWesi Indies Central and Southern Africa hm w Tumor Viruses Latent Life Cycle Virus Cell Integration usually Transformation Virusspecific proteins expressed No mature virus Changes in the properties of host cell TRANSFORMATION Tumor Viruses Transformation Loss of growth control Ability to form tumors viral genes interfere with control of cell replication TRANSFORMATION Both DNA and RNA tumor viruses can transform cells Integration occurs usually Similar mechanisms VIRAL TRANSFORMATION The changes in the biological functions of a cell that result from REGULATION ofthe cell s metabolism by viral genes and that confer on the infected cell certain properties characteristic of NEOPLASIA These changes often result from the integration of the viral genome into the host cell DNA TRANSFORMATION Among the many altered properties of the TRANSFORMED CELL are Loss of growth control loss of contact inhibition in cultured cells Tumor formation Mobility Reduced adhesion Transformed cells frequently exhibit chromosomal aberrations 33 Two Major Classes of Tumor Viruses DNA Tumor Viruses DNA viral genome DNAdependent 9 DNA polymerase 1 HltSt RNA Host or viral P0 ymerase Viral mRNA l Viral protein RNA TumorViruses WM m o ReverselranscriplaseVirusencoded VMWMWWWM lMPORTANT 39 A39dependenl RNA polymerase Host Vm A I I RNA pol II P mmwm N DN nmm Impo 39 RN rAm imok S icing Hoslsplicing enzymes HoisNA olymerasell DNA genome p I mRNA I Hoslenzymes olein p WUS ORTRANSFORMATION Intransformalion usually only EARLY functions are expressed J0 DNA Tumor Viruses In Human Cancer Papilloma Wruses cause natural cancers in animals cause benign warts ubiquitous epitheliotropic most human tumors are malignancies of epithelial cells 37 DNA Tumor Viruses ln Human Cancer Papilloma Wruses epidermodysplasia verruciformis wart malignant squamous cell carcinoma DNA Tumor Viruses In Human Cancer verruciformis Papilloma virus DNA Tumor Viruses In Human Cancer Papilloma Wruses urogenital cancer wart malignant squamous cell carcinoma Fapllloma vlruses arefound m 51 ofwomen wm cervlcal cancer Squamous cell carcinoma La nx Esophagus All histologically similar Lung 10 of human cancers may be HPV DNA Tumor Viruses In Human Cancer Papilloma Viruses 51 types identi ed most common are types 6 and 11 most cervical vulvar and penile cancers are ASSOCIATED with types 16 and 18 70 of penile cancers EPIDEMIOLOGIAL STUDIES BUT HPV 16 and HPV 18 do transform human keratinocytes NA Tumor Viruses in Human Cancer Polyoma Viruses Simian virus 40 juvenile hamster sarcomas transformation Polyoma mouse leukemia in vitro transformation Human polyomas JC and BK monkey sarcoma transformation PROGRESSIVE MUL TIFOCAL LEUKOENCEPHALOPA THY Polyoma virus transforms cells when the genome is incomplete Early functions are necessary DNA Tumor Viruses in Human Cancer Adenoviruses Highly oncogenic in animals Only part of virus integrated Always the same part Early functions E1A region 2 T antigens E1B region 1 T antigen E1A and E1B Onco enes Some very tumorigenic in animals EpsteinBarr Virus Burkitt s Lymphoma Nasopharyngeal cancer Infectious mononucleosis Transforms human D in vitro Considerable evidence for role in human caner Viral DNA found in small proportion of tumor cells hit and run DNA Tumor Viruses In Human Cancer Hepatitis B Virus DNA genome gnuApnlymgmu Reverse Irzns criplase V DNA DNA Tumor Viruses In Human Cancer Hepaims a cummued Vasl public health prnhlem Lnng latency DNA Tumor Viruses In Human Cancer Hepatitis B continued Epidemiology Strong correlation between HBV and hepatocellular carcinoma China 500000 1 million new cases of hepatocellular carcinoma per year Taiwan Relative risk of getting HCC is 217 x risk of noncarriers w a GunaX MEMSS use Gena DNA virus oncoproteins Large T ag Small T ag PP2A E6 p53 E7 pr E1A pr E1 B 55K P53 LMP1 TRAFs RNA Tumor Viruses RNA Genome Retroviruses RNAdependent DNA Polymerase encoded by virus REVERSE TRANSCRIPTASE RNA genome Reverse transcriptase DNA genome Integrase Integrates Host RNA polymerase II 1M RNA genome RNA Tumor Viruses RNA Tumor Viruses A normal retrovirus has 3 genes GAG internal proteins ENV Envelope glycoproteins POL Enzymes Reverse transcriptase Integ rase Protease RNA Tumor Viruses env Suliuce Glysopmlem su spun Yronsmembrane Givcopvolein m aw Mamx F El m gag cmm CA Care shell 24 RNA 2 moleculns val Wovens m av Pol mars in was WegmxelNVSZ RNA Tumor Viruses Groups of Retroviruses Oncovirinae MI Tumor viruses and similar Lentiviruses 4mm Long latent period Progressive chronic disease Visna HIV Spumavirinae 5 27 RNA Tumor Viruses Retroviruses known to cause human cancer Human T cell lymphotropic virus 1 HTLV1 Adult T cell leukemia Sezary Tcell leukemia Africa Caribbean Some Japanese Islands Human T cell lymphotropic virus 2 HTLV2 Hairy cell leukemia HIV 55 RNA Tumor Viruses Parental RNA Reverse transcriptase RNANA Hybrid Reverse transcriptase Linear DNADNA duplex Circular Duplex DNA Integration Re lication DNA genome in cell Transcription 5 Viral RNA genome mRNA protseein 28 Some retroviruses have an extra gene typical retrovirus R U5 GAG POL ENV U3 R Rous Sarcoma Virus R U5 GAG POL ENV SRC U3 R 57 Some retroviruses have an oncogene instead of their regniar genes Avian Myeloblastosis Virus R U5 GAG POL MYB U3 R Feline Sarcoma Virus FSV R U5 dGAG FMS dENV U3 R Avian Myelocytoma Virus M029 R U5 dGAG MYC dENV U3 R 29 RNA Tumor Viruses Viral Oncogene Vonc Cellular Protooncogene Conc RNA Tumor Viruses Protooncogene A cellular host gene that is homologous with a similar gene that is found in a transforming virus A cellular oncogene can only induce transformation after mutation some other change in the cell s genome 30 RNA Tumor Viruses What do oncogenes encode Proteins that are involved in growth control and 55 differentiation KINASE Growth factors Em musmlmm Growth factor receptors at Signal transduction proteins Transcription factors Protooncogenes and Cellsignaling gtSecreted proteins gtTransmembrane receptors gtGTPbinding proteins gtProtein kinases gtGene regulatory proteins 31 RNA Tumor Viruses The discovery of the acutely transforming retroviruses that contain voncs explains how cancers may arise as a result of infection These viruses cause rapid cancer in animals in the laboratory RNA Tumor Viruses In contrast Chronically transforming retroviruses cause tumors inefficiently after prolonged period of time Avian Leukosis Virus causes lymphomas R U5 GAG POL ENV U3 R No oncogene How does it cause a tumor 32 RNA Tumor Viruses ALV can integrate into the host cell genome at MANY locations but in tumor it is always at the SAME site or restricted number of sites Suggest tumor arose from one cell Something must be important about this site for transformation Crucial event must be rare RNA Tumor Viruses What is special about this site Myelocytoma tumors from several birds all have the oncogene close to this site It is close to Cmyc Oncogenesis by promotor insertion 33 RNA Tumor Viruses Genes can be quot assignedto sites on E E specific I myb mos chromosomes 5 a quotwe 1 2 3 A 5 7 E u 0 1 12 mosandmyc 9 E chromosomeB E Equot E E E a g g feschromosome15 21 2 X V 1a 15 1s 17 13 19 20 2 u 67 Cancers often resullt from gene translocations 39tt s Lymphoma translocation E E 3 EE a quot 13141515171319202122x Y 11 Oncogenesis by rearrangement Tumor Burkitt s lymphoma T cell chronic lymphocytic leukemia Bcell chronic lymphocytic leukemia T cell chronic lymphocytic leukemia conc myc a bcI1 bcI2 tcl1 new promotor lg heavy 8 to 14 lg light 8 to 2 T cell receptor 8 to 14 lg heavy 11 to 14 lg heavy 18 to 14 T cell receptor 14 inversion The multistep nature of Cancer mm mm Eugen1w Vm unmask fawn 1m Ixll mm wk nywn nylllon Herpesviruses Who here has Herpes Herpesviruses Herpes from the Greeks erpein to creep HerpesvirusesGeneral Herpesviruses are quite ubiquitous in nature 39 dnon mammalian extebrates Dogs cats bovine equine onkeys Sealions and ofcourse humans Chickms Turkeys Cat sh Salmonids Snakes Turtles ALLIGATORS Invertebmte hosts Oysters shrimp uitous ARE PREVEL T Herpesviruses are not only ubiq HE AN Presents problems with immunoeompromised patients 0rgan Transplant Herpesviridae HerpesvirusesGeneral Properties of the Herpesvirus Family Large Enveloped DNA Viruses Icosahedral Capsids DNA replication in host cell nucleus Herpes infections associated with severe disease in neonates and immunooompromised individuals dae Herpes Collect ALL 8 GOT HERPES Herpesviridae Arrangement ofthe herpesvirus genome Double stranded DNA genome rLinear genome invirion DNA circularizes upon entry into nucleus or cell Herpesviridae have genomes that rRange in size rrom 1507230ka relatively large rLarge encoding capadty 707200 gene products Enveloped Virus Size 1507200nm Icosahedral capsid ds DNA Genome Approx 100 genes a b UL l a c Us ta A lII ll TR IR 13 TR Structure ol HSVrl Virion Disease Herpes Virus 1 quot 39 HSV l Lytic Lifecycle Entiy Transport to nucleus Coordinate Viral Protein Expression And Replication Capsid Assembly Virion Maturation And E gress 1 Primary inl39ec onLYTIC INFECTION rAbsence of detectable antibodies 2 Latent infection er primary infection Viral DNA persists in target cell usually an immunopriviledged site 3 Recurrent infection Recrudescence Reactl39vation of latent genome Symptoms occurring at site of initial infection HSV l Lytic Lifecycle Entry Coordinate Viral Protein Expression Capsid Assembly Virion Maturation And E gres Herpesv 6 Heme im 39 me E7 S tab Atmhmzm m4 A7 Laws Auncm m mewwm mm quotmm mm m C7 Penmmn mm mm o D Immm Capndmd39lz udm HSVrl Lyuc L ifecycle Em hm 0mm Rzpl39mzl39nn Czpu39l Assme Virizn leun nn And Ing 1mm m mm Candilat Vin in mm Herpesvi u e nvelnpmmDerenvelnymzn svirus Mo ho enesis andE r Vanmin Impn Her esvlrldae g H me im Mnmhnvenei andEgress EmbpmqDumhpm a o r 39 Vanmin Impnn Herpesviridae Mammm Analysis Herpesvirus Egress and Plaque Morphology wildtype Synqtjal Herpesviridae a Herpesviridae E What is the difference between true love and herpes BY ERNVE BUSHMILLE Herpes lasts forever Herpes is a Love for Life Herpeva dae Herpesviridae Hernes Virus 1 Disease Hernes Virus 1 39 Disease Tropism herpes viruses canbe broadl 1 anary mfec39wn subdividedinw two subgroups based ontheir mum of detectable anhbodm use ofeither neurons leukocytes as the z Latent infection latent site oflnfectlon eAl ter primary infection eViral DNA persists in mget cdl N uronal herpesviruses Leukom39c hapesviruses eusually an imrnunopriviledged sitt The alphaherpesvlmses Beta and gm herpeswmses 3 Reeuuent mfeeuon ecmdescence HSVl H v2 MV monocyt s Reactivation of latent gmome EBV B cells s toms occurring at site ofinitial VZV 1 V 113 leukocytes infec on HHV7 leukocytes HHVS leukocytes Herpesviridae Herpes Disease Herpes Virus 1 39 39 Disease HSVl I atent Infection 1 Virus transported from Tropis herpes s can be broadly subdividedinw two subgroups based on their use ofeithermurons ot leukocytes as the latent site ofinfection Leukom39c hapesviruses Beta and gamma herpesviruses Neuronal herpesviruses The alphzherpesvimses HSVl H VZ VZV PV TRlGEMlNAL NERVE initial site of infection back to trigerninal ganglia 2 Asymptomatic No virus or virion proteins produced 3 Viral DNA resides in sensory cells of Trigernjnal nerve ganglion Herpesviridae Herpesviridae Establishment of L atency productive motion a Reacnvatlan m ex ressun impurmntO Pmduzova 0 mteenm x a n o39 o a z a of Sensmyneumn lt o Estzhhshmen u atentv v2 k HSV gencme msmne essseretea Epimm mt expressed m ch rem some Wat qenumes m m V ee 0 cps we sutgf eeed 3 i synapse II Latent Phase Hm Jrao rmmpmt minimumquot Herpesviridae E Establishment of Latency Pmduclwe lmwllun I HSV senemehrsmne esseeretee Ewmme Lens expressed ems ed Jens quotE gt e 1senswnepm lt n 39 smhushment some wa geneme gene ms through svnause Latent Phase Mun phase Herpesviridae E Establishment of L atency vmuctwe lmemlan 0 Re tiveuan m expressun mpurmnt Establishment of Latency Pmduclwe lma lnn I HSV sencnemsene assuaatzdepisome m s Expressed Pmdumw mCNS LEEng t K I39 i 0quota J i 39 quot 39 quotE me Esmhhshmgt TRANSPORT OF CAPSID u atenzy precpepte Pmdutnve semewn emes Semevira emes e gemcps irnn gigggged Q We cemepsirw xed 3 aquot svnavse 5 Evnapse 5quot Laieniphase Hmjegg39 LatentPhase Hmjcg mum tmmmrtr Acmepnsse namepnsse Herpesviridae r Herpesviridae Maimm m Emmy Disease Herpes Virus Patho enesis The nznrnn is an immunnpriviiegerl site mnne wasinn tactic nl39 virus Viral DNA exists in its genpnpic pm as an zpisnmewithin the cell may Lytic cycle genes are transcriptinnaily and functinnaily quiescent ADDITIONAL nvuvaE EVAme TACTIC ONLY LAT Is EXPRESSED 1 Primary infection Absenee of deteetable antibodies 2 Latent infection After primary infection rViral DNA persists in target cell susually an immunopnviledged site 3 Recment inrectipn stcnIrlescence sRezctivztinn nl39 istent genpnpe sSympt ms pccming at site at initial iniectipn Herpesviridae Herpesviridae Disease TRiGEM NAL NERVE HSVl Recurrent Infection 1 Upon stressor virus replicates and travels down sensory nerve ber to infect epithelial cells around the nose and mouth 2 Symptoms are usually a milder form of primary infection Herpesviridae Real vallnn m expressan imwrant HSV genuine nismne associaizd aplsame s expressed n o z H 39oKJ i v chimney v W 5 anagrams xx 0 quotin n synapse i n Latent Phasa Hm j i umnumty Acute phase Herpesviridae E Reactivation rrnm Latency sensury neuruns must survive repeated beuts ni iytic reactivatinn witheut compromising function A iimited number at sensnry neurnns may prnduce nniy a iew iniectinus viriuns tn iimit the nnrmaiiy vegetative iytic rep iicative lifecycle Additinnaiiy Hsv 5p ecil39ies antirzp uptusis genes such as 1c1gt345 which may ac to pretect neurnnai ceii death Disease A 39 39e 39e I atent Infections l HSVl site of latency trigeminal ganglia 2 HSV2 site of latency sacml ganglia 3 VZV site of latencydorsal root ganglia Herpesviridae E BY ERNIE BUSHMVLLE i must a Herpes is forever HERPES GOES LATENT and is PERS STANT Herpesviridae Herpesviridae E Clinical Manifestations of Herpes Infections Herpesviridae Disease l Gain access to body via mucosal surfaces or skin breaks 2 Usually cold sores sore throat fever and rarely Encephalitis 3 Less frequently found as a genital infection H SVl Recurrent Infections Disease Early stage 39 Over 175 million Americans 70 seropositive gt50 acquire before the age 0 39 33ofHSV1 seropositive individuals reactivate 7 5 have one per month 7 34 have recurrences between 211 months 7 61have recurrences ofl or less per year 39 Most commonly asymptomatic gt80 don t even know they are infected J A Herpesviridae a Herpesviridae m Disease HSV Z Primarv Infection Disease SV Z Puma Infec 10H 9 ONAL 1 Usually vesicular eruptions on the genitalia N Less frequently found as Herpes Labialis cold sores F Spread by sexual contact Affects both sexes Once was associated with b MAL U cervical carcinoma Herpesviridae m I erpesviridae a Disease HSV2 A quotV quot 39 Disease SV 2I m lent Infection 1 Viral DNA resides in sensory cells of sacral ganglia Heterosexual men Heterosexual women 2 No Virus or Virion proteins 0 l partner 0 l partner lt10 produced 0 210 partners 20 0 210 partners 40 11 50 partners 35 11 50 partners 62 3 Mllder 011th le 1 0 gt50 partners gt70 0 gt50 partners gt80 same locanon Ofmma infection within the genital area Her esviridae Disease HSV 2 Recurrent Infection Disease HSV1 amp 2 m p 4 of 9 adulw are infected with HSV2 Herpes encephalitis over 50 million Americans HSVl I Frequencies of recurrent infections 530 yearsold a 33 ofpatients gt 89 per year Reactivation from trigeminal ganglion r 33 0fpa1ients47 per year Hsv2 r 33 ofpatients 23 per year Neonatal infection I Less severePreSenmmn than Herpes meningitis HSV2 I HSVl genital infections less likely to recurreactivate Her esviridae Disease HSVl amp 2 quot 39 39 39 F 39 39 Disease VZV Primarv Infection Neonatal Infections C ken PoxVaricella 1 Infection occurs in seasonal epidemics as chicken pox Varicella 0 In utero 5 of neonatal infections 7 HSVZ 7 Ascending or transplacental 2 Contracted from another 0 Intrapartum 7580 of neonatal infected individual highly infections communicable90 POSHHtal transmissible HSV1 3 Systemic infection resulting in a generalized vesicular rash Herpesviridae Disease VZV Latent Infection l Asymptomatic with no virus or virion proteins produced Viral DNA resides in the cells of dorsal root ganglia N Disease VZV Recurrent Infection ShinglesZosler 1 Virus travels down the sensory nerve ber and infects epithelial cells innervated by the ber Herpesvin39dae Disease VZV Recurrent Infection ShinglesZester l Infections are unilateral painful vesicular eruptions localized to the dermatome usually in the head or upper trunk N Severe systemic infections are observed in immune suppressed individuals Her esviridae m VZV Recurrent Infection Disease Fast hapetic neuralgia Vay painful Lastsfm39 mnnthsa a Znsla39 resulves Dues not respond in Antiviral treatments Likely due in nerve dang from 1am ShinglesZosler He esvin39dae Disease Therapeutics for Treatment of Herpes Infections Acyclovirrelated antiviral drugs acyclovir famcyclovir valacyclovir gancyclovir iacyclic analogs of guanosine iterminate DNA synthesis all are inactive prodrugs Heiesvin dae Acyclovirarelated antiviral drugs acyciuvir alncyclnvir ydnvingancychvir Herpes Virus Treatment Questions Herpes Virus Treatment Acyclovirarelated antiviral drugs aquamarinch y 39 gancyclnvir acyclovir famcyclovir valacyclovir HSVrl ancyclovir similar drug but is active against CMV Herpesviridae Herpesviridae If Family 3 Type Species Hosts Adenovrridae 39 39 391 39 39 s Vertebrates A Fowl adenov Vertebrates Adenoviruses WWW o qmammmmmpggyamwm WMmmmem v Mcwon gtlt mum is m WHO S Emma Chime prmmitzb 39 KNEW M mmmf w w39 whim m J Mm 1 lm saw minim 1 wm mrxm Iliumw and mmtam Adenovims Proteins Capsid praKeins mm w lt vu com n DNA omen lt m I ma yum V v l W vanjL Nuclear pore cytopla smj Pentnns rupture phzgncytntic vesicle M 39J gt39 mmum 39 Mam 0021 17m ng n ac a Jig WML 11mm 5 ems d s a acaco 3 day m ied mm mfg W mu 3 I e mwmmgmg lime WWM Magma 7 I h w rm39m H v39rm n mm mm W H WM 7 Wm mm mem WW I I M I H mm mm mmm H mm mm m u w n w u u Sigmm l ditgp mmm L11T 1mzal mm 1 quot 1 9 Amwmg 51 m mail g MM 343 itamglm aux51 le imp ag WJigwlam b0 Wm WM fcmmg mim m 3m DNA gym m gu 9 3 SQ39V 7 3 D V quota 39 1 E E W 333 Wail by ME Lm m umg Sham digp aaemem TL Hm Im rglkm a Tam hm gmm mdQTP DNA pmbmmy mama blank 39 5 and mi 2513 mam 321 DNA L 3 CODE 9f GEE a m a prim WENA a NEW 23mm 3 vmva mw 1m tam M f QM T1 13 biap wacdl by 513 mm if Em mail Wham Mb 9 k 3g m LU lhitaifib f nl39v fa Ltrm lm g FrT s l Adenovirus Gene Elpression Phase GenesTranscribed Immedlale early EIA Early E1BEZAEZBE3 EA some Vlrlon protelns Late Late genes mostly Vlrlon protelns Adenovirus Transcription Map All transcription dependent on E1A Late transcripts have common 539 end 39 Eight transcription units Promoter Structure gt10 gt10quot 7 x 102 5 x i02 20 35 2 Dwtant raguiatory anal reguiatnry Core promoter upan sequunces Stlancprc ilt gti Pr umouzr at ranscriptional control region WWII 5 13pm A We I a ma mmmmmmhmmwm dimmgmm aiwmmsumimvmm x wank ame y AN EXAMPLE OF PROCESSING pnmwmsmw c mammm pawamyme mw wrunremuval mm h mm pawn m m quotAW Adenovirus Transcripts are Spliced m mum mmutoocculW M39s Nuclear RNAs gt mFtNA hnFlNAs have 5 caps and 339 polyA L RNAs seem to map to e same promoter How to get small RNAs 5 terminal tripartite leader and body x c A x Cample mummtmn by mm Rm mutt Nsxm Mxm Campquot mmmmhym What is E1A N p300 An Adenovirus IE protein activates all Ad promoters Doesn t bind DNA Nonspecific TA pol 1 amp Ill promoters Only sees Ad promoters in infected cells How Does E1A Work E2F release from pocket proteins Pathogenesis mm Adammizmm DRE Y 39 g Amtquot WWW my mama immamg w mmim F San 39 my s E mam m aw1mm gammy E m mmmm um mm Limme mums M Vaccines 0 MW mmmmw mg m m e eamated W 9 m bngi g w j V ME 3 m am mm m i i Wm gwi W 333 am f WEI 111 WWW m f a mmami mima 533mm La ZaWSW whamawwi dwi hmm g 39E39G L O 8 11 39 111ml III 110111311110 IIlOlllIu p111 110111311110 11111 IO 391 1mm 11 31st sonpmd 01 quL 13N 3813 V39IEI Hepatitis B virus Genome partial dsDNA Family Hepadnaviridae Related Viruses Duck Hepatitis B Virus Ground Squirrel Hepatitis Virus Snow Goose Hepatitis B Virus Woodchuck Hepatitis Virus Wooley Monkey Hepatitis Virus Incubation Period 30 180 days Transmission Blood and Sexual Contact Acute Attack Mild or Severe Serum Diagnosis antiHBc antiHBs or HBsAg Prevention Vaccination with recombinant hepatitis B surface antigens Hepatitis B lmmunoglobulin History of the Hepatitis B Virus Viral hepatitis is the term reserved for infections of the liver by one or more of different hepatitis viruses The terms hepatitis A and hepatitis B were first introduced by MacCallum in 1947 in order to categorize infectious epidemic and serum hepatitis In 1963 Blumberg discovered a previously unknown protein in the blood of an Australian aborigineThis protein was denoted as the Australia Au antigen By 1968 other investigators had established that the Au antigen now known as the hepatitis B surface antigen was only found in the serum of type B hepatitis infected patients In 1973 Dane found viruslike particles in the serum of patients suffering from type B hepatitis These particles were designated as the hepatitis B virus HBV HBV has a world wide distribution and is of great medical importance because it may be the most common cause of chronic liver disease including hepatocarcinoma in man It is probably the most important human carcinogen next to tobacco There are more than 750000 carriers in the US and more than 300000000 world wide of whom a quarter are in Asia About 10 of the population of Asia are carriers in the US about 1 The virus is spread mostly from mother to child or can be transmitted in saliva or semen it is thus also a sexually transmitted disease and is now also associated with AIDS The epidemiology ofthis virus is similarto that of AIDS except the virus is much more infectious Many carriers do not have infectious symptoms but can still transmit the virus Three closely related viruses have been found in lower animals and have been used as model systems these are the woodchuck HBVWHV ground squirrel HBV GSHV 1mm and the Pekin Duck DHBV These viruses are now 3 33 v classified as hepadnavirus These viruses have as yet not been propagated in TCand thus many details oftheir mama replication are unknownDifferent viruses cause hepatitis mart1g m These are HAV a picornavirus HBV which we shall 511E100 concentrate on during this lecture Hepatitis C which is a quot flavivirus and delta agent which appears to be a defective r virus or viroid that requires HBV for growth amine lilllillrrr L sirmil Hepatitis B Particle Types The hepatitis B virion also known as the Dane particle has a diameter around 42nm The outer envelope contains high amounts of hepatitis B surface proteins The envelope surrounds the inner nucleocapsid which is comprised of 180 hepatitis B core proteins arranged in an icosahedral 3mg an arrangement The nucleocapsid also contains at least one detergent hepatitis B polymerase protein as well as the HBV genome 8911M HF mlem m core a Two other subviral particles can be found in an infected individual39s serum namely the hepatitis B filament and hepatitis B sphere Both particles have a diameter of 22nm and are composed solely of hepatitis B surface proteins The sphere is composed ofthe small and middle hepatitis B surface proteins whereas the filament also includes the large hepatitis B surface protein High levels of these noninfectious particles can be found during the acute phase of infection It is possible that the presence of such high levels of noninfectious particles may allow the infectious viral particles to traverse the blood stream undetected by neutralizing antibodies Core Particles Purt ed vtrtons possess the HBO protem Whtch aggregates to form the core parttote Thts H v turn Geom etrj39 I a n 7 15 V t u n ww r z o a 9 m3 a hegatms B Surface gro ems The Hepatitis B Genome tn vtnons the genome ts ctrcutar yet onty pamaHy doubterstranded The genome ts approxtma et 3200 nuoteottdes H v tength HEW vtrtons oontam both DNA and RNA Moreover some regtons otthe packaged genome can be d ordouote stranded T ere are fourdeftned overtap mg open readmg frames ORFS m the genome Whtch resutt ton of the seven dtfferent hepatttts B protetns The genome atso oontams genettc etements Whtch regutatetevets oftranscrtptton detenmne the s e tp r ad n ran n r r the nuoteooapst The four ORFS tead to the transcnptton and transtatton of seven dtfferent HEW protetns through use ofvarytng tnrframe start oodons ORF P occuptes the matonty otthe genome and encodes for the hepatttts polymerase prot m F s encodes the 39 ORF quot ro ein ORF x encodes the hepatttts B x protein 39 than t r hat rephcatton to occurThe four ORFS Ira crtptton are controHed by fourpromotereiements prest preSZ c e and X and nts ENH AH HEW transonpts share a common adenyt t n stgnat Resuttmg transonpts range from 3 5 e oorepregenome promoter oontrots the transcrtptton otmutttpte RNAS Whtch have 5 osome soanmng otthe pgRNAtranscrtpt F 8 contains three H39Hrame start Sites which direct T ere are two promoter t Whtch otthese protems namet th st p d pr moter T e prest promoter oontrots transcrtptton of n s RNA ts the epsttonrstem toop estem toop The regton ehsspsTesTTuh ssweu as YEVEYSE TTshssTTpTTeh enhe HEV pgR NA The Live Cycle nnhe He all a s v m as uVaHuthEqusEs ThusuTTsT snssh spesTTTssTTy END a new sspshTe eT suppuTTThgTTsTepheaTTuh TheughThe WWW Mama mn ReTease TT eTTsThe mus e ecwe seTTTypeTeT 4 TepTTssTThg EV uthevemahepaucsnes heme have been Teuhe m he shTe m suppun TTahscTTmTah sen TThes sTe sspshTe eT suppumng Vwa mGms The ava abmty enhese sen sysTems has aHuWed TeTThe eTusTesTTeh eT mush enhe hepsst a TTTesysTe 39 V A avuncua mg tha nudEucapsTd Ts behaved m he TTshsTmTTee m The husTesT methshe The HEV geheme uhseshhg DEEUYS snhe husTesT methshe HBV Genome Repairand Transcription mmquot m The HEV DNA Ts bvuugm mm The husTeus e TT Ts YEpaWEd m The cave eumpT TT uT he dsDN mm TemuvaTe The WW 5 T T T TusmTessh T T hmhe TmTymeTssepTeTeThTssweus evsT TTgsTTeh enhe shahes ohseT e geheme Ts mm TeeTTeuTaTTzee eTThaTTeeTaTTe TummeT W s aE NmES s13quot pmducmg The VaHDuS HEV TTaTTseTTstTeeuTTeeTeTHBvTTTmeTTT syTTThesTs mum a rw H she pgRNA gEnEYaHDn TheTTshs mums I geheTaTee can he dTdeed Thm me eaTeguTTes N H subgenumm she genumm The smaHev mm x W 3 e a e s T h p TsTgeT genumm TTshssTTst sTe TehgeT Thsh ehe 4A geheme Th TehgTh and serve m pvuduce e we and pnlymerase meeThs ApsTTTsuTsT genumm UanSEHpL TashThg The ATG 513quot BEIan TeTThe e pYD Em Ts eesTghsTee The pgR NA SmEE TT Ts sheseh spesThssuy by The HEV TmTymeTsse pTeTeTh TeT packagmg 39 m NucleocapsTuAssemth aTheThg enhe TmTymeTsse pTeTeTh m The rs emrmup Tesuhs Th pgR NAEncapsTdaHun H Ts behevedthanhe mammary wmamwmmmwm mum mm m ammo Temps sTe packaged by seTe pTeTeThs Thm WHEIVVS The hTTT HM T quotD4 T T Th TTTh h h Th Tsupyumm TsTessTeehesnhea transci iiii i i di ua eDNA strand therstemiuup as itsteiiiyiai i i Eriptmn in i di i i enaii iibed ndeieetides in the DRi regiun As such this segment er DNA ean be transferred m the 3 DRi seddenee and reyerse HUN NFFHN VHF HP PHI HPH pgRNA iernpiaie is degraded by RNase H adiyity ufthe imiyrnerase prutein Huvvever the is In id eapped diigdnbdndeiemides aune 5 end ufthe RN 7 DN prirnerreir DNA strand synthesis Tne neyyiy generated RNA nrner is rneiyed and baserpaired m the 5 DRZ regiun an the 7 DNA sirarid Dnee transiucated DNA S Liarid synthesis begins and EDNUHUES tn the end ufthe 5 end ufthe VJDNA sirarid Tu M d n HaH ii DNAsra ae punieir n M d eDNA sirarid TypicaHy the DN in packaged nepadnayirai DNA Hepatitis 539 Core and e Proteins The nepamisa edre prutein HEE isthe maid Features of HBc Protems 8 Gene nmpunent ufthe ndeieeeapsid sneii packaging the HEN genume Tnis ids arninei aeid prutein is expressed in the mpiasrn er infected eeiis Protein I Pnc an arged arninei aeids Tne regiun d H e Ssuciated With ndeieeieapsid assernbiy iies thin the rst ma arninei aeids Tne CrtErminai mien appeartu be inyeiiyed in ndeieeiiide packaging HE is abie tn rdrrn panieies With uutvvard puinting spikes eyen ir a rdreign peptide seddenee is spiieed inm the HE ending re iein As Such HE ean be used reir dispiaying rdieign prutem deirnains an the sdnaee eir HEN ndeiedeapsids Unhke the edre prutein the hepatitis E e antigen nas a different fate Tne e eaii igen i despite me One inediy HEVrprudumng eeiis Hign ieyeis eir HE eaiiiei persis tentWHV WEEUEIH ir a HEErminuS variant is used reir WEEUEIH Features of H31 Protem 8 Gene Hepatitis B Polymerase Protein Tne argesi ORF in the HEN endrne Encudesfurthe nepamis pm E pdiyrnerase prutem HEp Tne prmein is an id in size and nas RNA and DNA de Endant pdiyrnerase activity Hap piays a key idie in HEN genume generatiun as WEH as pgRNA Encapsidatiun HE is packag tugetherwith pgRNA Wiinin HEN ndeieeieapsids HEp nas been diyided inm rddr haramerized deirnains Hepmis B surface Pmei Features of H35 Proteins 6 Gene VWhiri the HEN genemei the regiein eneeiding the HEN surtaee breteins ebntains three ineframe start sites Which share a cummun terminatiun ED Em Because at this the variuus HEN surtaee brbteins are aii reiated tei eaeh either by a shared regiein Wm rinvvri as the Sedumain 39 quot rim riieim iiiae e EQHEAKE Protein i ms mi LHEs Small Hepatitis a SurfaceAntigen HEsAg or SHEsAg This bretein is the smaiiest eitthe hepatitis E surtaee pruteins eentaining suieiythe Seduma it aisei has been reterred tei as theAustraIia antigen Hg fuuretransmembrane sbanning regieins The bretein is the prime eenstituent er aii hegatitis a ganicle terms As such this bretein abbearste e manutaetured by the Virus in high guantities it aisei eentains a highiy antigenie ebitebe Anaiysis eitthis ebitebe aHuvvs terthe sub in eit HEN earriers Middle Hepatitis E Surface Antigen MHEsA This intermediate br midd ErSiZEd HEN surface brutein ebntains an additibnai 55 amin ramd Extraceiiuiariy Large Hepatitis a SurfaceAntigen LHEsAg This bretein isthe iargest er the HEN surtaee pruteins eentaining the Preesi demain asWEH as the PreeSZ and s demains The Preesi dumain s seguenee abbearste be HEN bretein inveivediniiverattaehment e Tee idema additienai giyeesyiatien site but eentains a myristyiatiein s terminus aneher he 7 errn e in en ignai at its Ne ingt Nt inusteith membrane Overaxpre ssibn uf iiiiaiiririiiiiiiiii e W tirst suggested that ER retentiein Was due tei a cytusuii taeteir binding the mm d by mesttei be the ne respunsibiefurmediatingvirai attaehment entei its heist eeiis Huvvever the reeebteir tbr HEN has net been iseiiated Hepatitis BXProtein The smaiiest ORF in the HEN geneme eneeides tbr it riinrtinn is unknuvvn iguing etteet HEX may have en HEN inteeted eeiis ere nre inn ufHEi HEX has been shewn tei bind entei bsai a brbminent eeii tumuuresuppressur bretein PATHDGENESIS is must emeientiy transmitted threugh bereutaneeus intreduetien Sexuai transmissiein and berinatai iiu HEN and biuud With iesser The high ieveis er rebiieatibn Thevirus amuuntsfuund in saiiva and semen The Liver The ttyer plays a ptyotat rote tn hetptng the body matntatn adequate teyets ot metabotttes tn the blood stream The tt or our blood stream On average the ttyer wetghs out tour pounds and ts the only organ able to regenerate ttsett eyen when up to 25 ot tt ts remoyed The ttyer has many tuncttons tnyotyed tn dtgestton and metabottte conyerston Some ot tts key tuncttons are Bile ProductionThts ttutd ts stored tn the gattbtaddertor release when needed tn dtgestton Btte ts necessary tor the dtgestton ot tats as tt dtssotyes tat own tnto smatt gtobutes Btte Works tn the same way as detergent tttts ott ott greasy ptates Btte atso asststs tn the absorptton ot tat sotubte yttamtns t e yttamtn A D P and tlt tn addtttort btte conyerts betacarotene to yttamtn A Glucose omeos asi B the torm ot gtycoge a potysacchartde when there ts an oyerabundance Gt cose can then be released trom the glycogen when needed b the bod rt u t the organsw I 62 Fat Metabolism The ttyer ts tnyotyed tn the synthests of damaged tatty actds trom amtno actds an sugarst tn the ener y Excess tood ts conyerted to tat tn the ttyerwhtch ts then transported to the adtpose tatty ttssues ot the body tor longerrterm storage tood tn ts detoxtfled by the ttyer The ttyer regulates protetn metabottsm The ttyer atso combtnestoxtc substances tnctudtng metabottc Waste rugs atcoh t t excreted trom the ktdrteys Thus to haye proper ttverfurtcttort you must haye proper ktdrtey tunctton When etther the ttyer or the it ttrtn h H1 CellITissue Tropism The prtmaty stte ot HEW repttcatton ts the ttyer However yartous extrahepattc sttes haye been proposed abnormattttes somettmes found tn pattents wtth acute hepatttts DIAGNOSIS 39 39 t I r t39 tnto tour stages t tncubatton pertod whtch ts the ttme between tntttat ytrat entry tnto the cell to ttrst day ot symptoms tt prodromat or prertctertc pertod ttt the tctertc phage and ty recoyety s m toms durtng the onset ot acute hepatttts B ytrat tntectton vary dependtng on the tndtytduat chttdren and some adutts tntected wtth the ytrus neyer show any dtscerntbte symptoms However most tntected tndtytduats expertertce a certatn teyet ottaundtce whtch tends to deyetop soon atter the ytrus can be detected tn the btoo ert taurtdtce ts preceded by mttd teyers fattgue matatse toss ot appettte and somettmes nausea and yomtttng Durtng the tctertc or blood borne hase an tntected tndtvtduat s urtnetends to haye a dark gotdertrbrowrt appearance Thts s often tottowed by the ng ot the stoot as well asthe yettowtng ot the Sktrt typtcatty seen tn taundtce uaundtce ts constdered cttntcatty apparent once totat btttrubtn teyets are greater than 274mgdt 0 Signs of Liver Trouble nr ht m H r n t att tt r problems are attrtbutabte to the Hepatitis Bytrus Hepatitis A c D E e ytruses as well as atcohot chemtcat bactertat and other condtttons can damage otte s ttyer The tottowtng ts a ttst ot the common symptoms ot ttyer problem Yellow dtscotoratton ot Sktrt andor eyes Abdomtnat swetttng or seyere abdomtnat paint Protonged ttchtng ot the Sktrt yety darkurtrte Pate stoots Passage ot btoody or tarrttke stools and Chrontcfattgue nausea or toss ot appettte tt one suspects one has a ttyer condttton tt Early ts htghty detectton may help stmpttty treatment Blood T l lul 39 39 quot vartety of viral infection av Botn HB and antiaHBs antibodies at a ieyei oti miUmL PCR nas aiso been used in detecting iow i biood and iiyertissue sampies iSA HBsAg proteins as iow as o 25 rigmL eyeis ot HEW DNA present in botn Reference Ranges in Clinical Biochemistry Tests AST w i i i i maini in heart kidney tiver muscie detec ed in tne biood vitamin B deticiency and pregnancy may actuaiiy decrease ieyeis ottnis enzyme tound in tne biood ALTM39 39 SGPT 39 39 39 Miii maini in tne tiver but can aiso be seen in iower amounts in heart muscie and otner tissues increased ieyeis ottnis enzyme in tne biood can be attributed to iiyer damage kidney infection cnemicai tokins or eyen a cardiac intarction neart attac 39 A Mi i i uiiuiiibone andiiyertissue rowng cniidren typicaiiy naye an increased ieyei ottnis enzyme in tneir biood compared to aduits for children 407400 UL iii i i uiiuiiiiiyer n iimntion tnisenzym diseasebi d t b t and drug abuseto nameatew lnH ll 57225 UL i i uiiu in brain neart kidney tiver iurigi and skeietai scien ii ceiid atn mainutrition oriow tissueorgan actiyity Albumin 35750 gL i Tnis protein is syntnesized in tne iiyer and is inyoiyed in maintaining biood protein base ieyeis tiyer damage may resuitin iow ieyeis ot wnen aibumin i i i i may ieakinto surrounding tissues resuiting in sweiiing known as edema Bilirubin non neonatal Totai conjugated and unconiugated M7 umoit i Biiirubin is a byproduct ot red biood ceii breakdown it is actuaiiy tormed wnen tne nemogiobin ring is opened tnrougn otner enzyme actiyities Biiirubin istypicaiiy iiverdi a m ri riit i i t i it due to sometypes ot drugs and nemoiytic anemia Bilirubin non neonatal Direct conjugated 075 umoit Sernlngical Pattems Obsewed During Acutc ILBV Infection CLINICAL TYPES AnnHE Acute Hepatitis The miidest attacks are m lt 3 y o a g 1 a 71 9 2 a g i 3 z 7 lt imal I 39l l39 Illa trom wnicn recoyery Deteuimi Letet is typicai tnrougn to tuiminant tatai yirai nepatitis iicteric attacks in aduits are marked by a V prodromai period typicaiiy tnree to tour days but H I 2 3 4 gt 6 7 X may iast up to two or tnree weeks during wnicn Mmquot pmmlmm patients teei sickiy sutter trom digestiye symtoms sea and may in tne iater stages naye miid pyrekia Otner common symptoms are rigors ioss ot desire to drink aiconoi or smoke m and occasionaiiy seyere neadacnes Tne prodromai period istoiiowed by tne 8 E m 1 2 ataiSei darkening ot urine and iigntening otteces toiiowed by tne deyeiopment otiaundice 39 39 u Tnese ekcessiye noduies and tibroids resuit in tne distortion ot tne normai iiyer arcnitecture and intertere witn bioodtiowtnrougn tne iiyer cirrnosis may aiso resuit in tne inabiiity ot tne iiyer to pertorm its tunctions as abnormaiities progressiyeiy deyeiop Fulminant Hepatitis This is a rare form ofthe disease which usually overwhelms the patient within ten days This form may develop so quickly that jaundice is inconspicuous and may be confused with acute psychosis or meningoencephalitis On the other hand the patient may become deeply jaundiced Forboding signs may be repeated vomiting fetor hepaticus confusion and drowsiness The 39flapping39 tremor may only be transient but rigidity is usual These are then supervened by coma indicating likely acute liver failure The patient39s temperature rises jaundice deepens and liver shrinks possibly accompanied widespread hemorrhages The levels of serum bilirubin and transaminase are poor prognostic indicators because transaminase levels may actually decrease as the patient39s clinical condition worsens Prothrombin is the best indicator of prognosis Frequency of the fulminant course varies depending upon the type of viral hepatitis and prevalence of hepatitis B carriage PostHepatitis Syndrome Adult patients feel lessthanperfect for variable spans oftime following acute hepatitis Typically this period lasts for a few weeks but may extend into months Common features are anxiety fatigue failure to regain weight anorexia alcohol intolerance and right upper abdominal discomfort The edges ofthe liver may be tender Serum transaminase levels may be up to three times that of normal Hepatic histology reveals mild residual portal zone cellularity and fibrosis and sometimes fatty changes in the liver cells These conditions do not differ from patients found to be recovering normally and who are now symptomfree Thus a liver biopsy should not be done within six months after acute hepatitis as there may be difficulty in distinguishing the residual changes from developing chronic hepatitis Relapses In 18 15 of cases some ofthe original attack is duplicated but typically in a milder form Often relapse is simply shown by an increase in serum transaminases and bilirubin Relapses may be a result of premature activity or intake of large amounts of alcohol Multiple relapses may occur However full recovery may follow though in some patients relapses may indicate progression to chronic hepatitis 39 I quot 39 f 39 HCC l 39 t quot Ilar carcinoma is the technical term for liver cancer This form of the disease develops after a long time in individuals suffering from chronic hepatitis B infection What events trigger the development ofthis disease are currently unknown TREATMENT Disclaimer Please consult with a professional health care provider BEFORE doing anything suggested or implied This information is solely provided for academic training purposes Treatment Acute Infection At present there are no specific treatments for benign acute viral hepatitis Therapeutic effectiveness of interferon use on the prognosis and course of acute HBV infection remain unknown Treatment Chronic Infection A number of elements has been used in the treatment of chronic HBV The goals oftreatment are threefold to eliminate infectivity and transmission of HBV to others to arrest the progression of liver disease and improve the clinical prognosis and to prevent the development of hepatocellular carcinoma HCC Currently there are several treatments being used Interferon alpha use is most common but now lamivudine STC and others are being looked at as potential therapeutic agents Combinations of antiviral drugs are being used with some success Liver Transplants Liver transplants are sometimes required by those with extensive liver damage due to viral or nonviral causes However there are many factors to consider before getting a liver transplant First and foremost transplantation is a complex operation that requires a suitable donor Also there is usually a long waiting list for those wishing to receive a transplant Moreover transplanting a liver into a chronically hepatitis B infected individual presents the likelihood that the newly transplanted liver may become reinfected Vaccination Currently the best method to prevent HBV infection is through vaccination The most common vaccine on the market is derived from a recombinant yeast source The small hepatitis B surface protein SHBs is generated by yeast cells Expression ofthis protein by yeast results in SHBs particle formation However particles are not secreted by the yeast Disruption of yeast cells is performed in order to liberate the produced spheres into solution These particles are then purified through clarification ultrafiltration chromatography and ultracentrifugation The purified particles are then adsorbed onto aluminum hydroxide to which thimerosal is added to preserve the solution The two yeastderived vaccines licensed in most countries are EngerixB SmithKline Beecham Philadelphia PA and Recombivax HB Merck amp Co West Point PA Both thankfully appear to be quite effective as vaccines allowing for immunization against the various forms of HBV There has been a recent surge in development of new hepatitis B vaccines made ofthe large and small hepatitis B surface proteins or DNA vaccines or even combination vaccines such as a vaccine providing dual protection against hepatitis A and B Twinrix SmithKline Beecham Biologicals a g much an39 tti nnDsmvdnpnlmt Disease Disease 11 We m mama 13M hm Mv m mm m m 52 mam 4 7mm im eim NEEmail RENAME nW wm WEBWWW Disease eexmw 11 la V 3 hm Disease I r Disease eV 1 si39r n i gemmm 353mgka len evaruses quot gs Vg W wmc v a fZV fie d iteiig roof ganglia Reac vatlon LAT expreseon ImuoI Ent HSV gmome memne assaCIa Ed eplsome T 5 ex res a Reac vatlnn LAT expresecn Impor ant HSV genome mscene assoclated apsgtme LAT IS a reegd tn CNS Pmaucuve Infection Sums Pmducuve Infectlcn sum 39 Senscrv neuron Sensory neuron Establishment Establl arlanency flaw some vlra genomes i t some vlral genomes gsxggscaw s mreu 9n supprgged gazetgsgws through a up Latent Phase Latent Phase Host Jegg Immunlty ACIJE phase Dmdmiive suremen Reaeuvauon a LAT Egttpre czn Impartznt Same viral genomes get to CNS through synapse latert Phase 9mm e an mm mg v bgag l 31a Minimum m We d m Wags 2m LEE mam 6W WW 533411 baggy mg I I mdhmd TL Agquot msg p quotc as Ree v m magnum 53 m m 1quot same we gemmes 533VE get to as mewsquot d suppress synapse laterk Phase Reactivation LAT expresson im penant HSI gmame msmne assxlated epuseme LAT VS expresse 7 Senser neumn lt Y Es zbllshment Some viral genemes r 3 gesggsius through suppregeg latent Phase ue na l 1e 35 Seen 1 L r isgnfzc a Px oduct a xxptoxxxatic e 39 939 7itl C B H y okur You 7 Pat ex t xc apt xxx ixxuxxux feltti o vnig h lt mm pmm 1 ow me a a sip m A J 39a EEDZPL P ng WEBW N H off l u39i l d xi exlare n1fecteltl b Lunwmwuun numbing a avs so It a W n a arm ammxwi w 1311 r m mam as LIB nos mm mt mtaw mazaimi g l aemm mm as icted nausea 331 he eore 3 W 0135 W Wm m ss sma a mxmnf T337 can Extv xiv31in mc wae j smbm 31m mm E m g W115 am may 3333 a a m m Gina WWm 3283 vil Ieve op 39u P oxne patients n c 31 ere equotentuall tllevllatievnt elie Lof 111Ilolroliferlti e sense or 1 39lljllollia VIPSJS VJ Kiampcagi gg Sea mcwmaw 81331 meg ESLEW 33 Emma 1mmm 621 N b 39 ai a 3 mg Kim mm mt mm 11me MEL3amp1 3 mm mam I pe VAULTUGJFJQ Kansas3 SarcomaAssoa39ated Herpesw39rus KSHV Lher herp uiruscs KSHV ls nut ublqultous Generally saraprevalance of KSIHV correlates KS incidence a primary disease was dcnnficd Prlmary Infecnon precedes lifelong latent Infection thslologlc signals haL serve to actlvale KSHV An Infected dIVIDUBIS are currenlly unknaivrl only a small percentage of me Infected Indlvlduals develop any dksease Risk of K5HV posmueHIVposl1kve Andlvlduals to develap KS wllhln 10 yrs is 100 Risk of KSHV posluveHlvvneganvc Indlvxduals to develop KS is usuuo 1120000 Immunulogzcaz status of an nrected Individual plays a full n the e omen a the disease H s KSHV repllta o and vice versa Inrects B cclls and endamenal cells DISEASE W S39 39 Italian ED onors 39L Jga Gyromes and Immune Regulators VIL 6 VIRFS Interferon regulatory factors VMIPS Macrophage Inhibitory Proteins VMIRS Modulators of Immune Recognition E3 Iigases that target MHC 1 on database screening a cellular homolog has been recenlly ldentl ed Goto e 3 IBC 2003 Potentla Transforming Genes Encoded by KSHI VWSMN 11 1M WHEN magma zz um Kllobases Reglons conserved wlth other herpesvlmsa 2 Regions uneque m KSHV and Rbadlnavruse VIRFI VIRFB vacI ln VGPCR ORF Kl Kaposin vBcl Z VFLIP quot vSurvivin v1L 6 Latency AssociatedNuclear Antigen 1 LANA1 Latency AssociatedNuclear Antigen 2 LANA 2 g e Inc 395 chn rn cterlsl lcs I uo naming c In he Whole Ar a ag ng Syslelns Ie Amman lg Systelns h mg 1 package tgmodi ed geP 39 ne inm Hsv aw A A w r V we Eegwmmf x2 PARVOVIRUSES The parvoviruses parvo meaning small are a group of very small DNA viruses that infect many species of animals Parvoviruses are divided into two groups The parvoviruses which require coinfection with adenovirus for their own replication constitute the genus dependovirus previously called the adenoassociated viruses AAVs These viruses do not cause disease in humans The second group of parvoviruses the genus Parvovirus do not require a helper virus like adenovirus for replication However they multiply only in cells that are in the process of replicating their own DNA The human autonomous parvovirus B19 virus replicates in erythroid precursor cells where it can produce aplastic crisis in predisposed individuals with underlying hemolytic anemia or immunodeficiency The parvovirus family contains two subfamilies with three generae each VIRUS FAMILY PARVOVIRIDAE SUBFAMILY GENERA PROPERTIES MEMBERS Parvovirinae associated with mainly warmblooded hosts Parvovirus mammalian V MVM minute V mice Erythrovirus human 819 Dependovirus dependent AAV2 Densovirinae associated with insect hosts Densovirus insect Junonia coenia dV lteravirus insect 80mbyx mori dV Contravirus insect Aedes aegypti dV Morphology Particles are icosahedral 1826nm diameter There are 3 capsid proteins VP13 VP3 is derived by protease cleavage of VP2 The capsid makes the virions resistant to inactivation by pH solvents or high temperatures 1h at 50 C Genome Parvovirus genome is linear monopartite ss DNA of approximately 5 kb in size Most of the packaged strands of DNA are minussense but the adenoassociated viruses package equal amounts of plus and minus sense DNA The genome also has palindromic sequences at both the 339 and 539 ends which form quothairpinquot 8 structures These quothairpinquot structures are 3355 criticalforgenome replication Parvovirus a w 30 20 m repI39Cat39on and assembly 0 C m the i m mugE i iriztwquot t m nUdeUS and 395 dependent Upon hog PeHUIar mwnn iicunning uuscni anquotAnmpntinnuwua enzymes The mechanism of replication of 3932ng 3390 2 m m 13920 the genome is peculiar and unique to the 15 9 virus family The hairpin structure at the 339 J end is used as a selfprimer to start gmquot 5 synthesis of a plussense DNA resulting in g double strandedDNA The hairpin structure is then again used as a primer to transcribe Egg 10 30 40 3 3 39 w quot1quotquotquot F EE EquotE lt f quot more mmus sense Strands from the ds DNA m i 333quot hauntHun rgna nnrl annualIn ESE 1m Ila l so Emu CUT III I 395 Replication The pathogenic human parvovirus 819 is difficult to grow in culture so comparatively little is known about its biology The receptor molecule for 819 is tetrahexosoceramide a glycolipid erythrocyte P antigen which explains the erythrocyte tropism of the 819 virus mm mm 2mm 0 mer nan mnsvnu nnowm w mm Emqu n mm om pumawa mm naeth q O Len Wm amp muse 0 mm uHuwm scheme abuve All pawnvirusvs are highly dependent nn veqm vnh n m w HDNAsymhesws muve dependent vequwe m1 22H machmevy mus he pekusmv vephcauun N h n mummy nu amen m m puuny undevsmud HUMAN VIRUSES and DISEASES Tne 8A1 VIYUS nas been aSSuciaIEd Wiin ineurnaieie arinniis received much aueniinn as nnssihie veeinrs Inr genelherzpy ADEIIoeASSogy ATED VIRUS Fium VIUS Muighuiugx 1988 enureniiii LivingSIune rnnIiH r M n n disease eryinerna inieenesurn E1 aIse knuwn aS In disease Fim vim Mergneiegj 1988 enureniiii LivingSIune Emir disease is a mild childhnnd iIIness chzrz erized hy aIaeiaI rash knnwn as quotSlapped cheekquot AIse seen are Iacerhke er IEIIcuIaIEd rasnes un Ine hunk and emernmes e a Huwe res en piuseveiaiweeks IeIIeWIng inieeiien Wiin Ine VIYUS In resnense In changeS In S11muh suen aS Iempeiaiuie Sunhghh and erneiienai siress ASympIumaII inieeiien win 819 nas been vepuned In abqu 28 mi eases thngenesis mm m7 stamng 1 my 4 m a Diseases Erymemx mwcnasvm am and pmmus Anma g a 2 Spread Fm oss Aplasuc cm Emmmd aplasna A lmIWUM V39mmu Tm mvu ves an acme depvessmn m m pmdummn u l 12d mum ceHshum m bane manuW dwec y due m Vmunnm mm L iwarex nummmu Mu Humlwm HmI varlllesymrwm gnu Imam Pm e w 12 m a 15m 2224 as 2a was First disease measles rubeola Second disease scarlet fever Third disease German measles rubella Fourth disease quotFilatovDukes39 diseasequot doubt exists over the existence of this as a disease entity Fifth disease erythema infectiosum slapped cheek syndrome Sixth disease roseola infantum herpesvirus 6 infection quotexanthem subitumquot ln 80 patients there is also arthropathy temporary arthritislike joint involvement particularly in adults 819 infection in pregnancy is also associated with early foetal loss although the probability ofthis appears to be low lt10 Recent reports suggest that AAV infection in human semen may contribute to male infertility although the evidence for this is still tentative Control There is no specific antiviral therapy orvaccine for 819 infection and most individuals do not require symptomatic therapy Patients with aplastic crisis require erythrocyte transfusion support until the bone marrow recovers Administration of normal human immunoglobulins to immunodeficient 819infected patients may produce transient amelioration of viremia and anemia Most persons infected with 819 virus are asymptomatic when viral shedding is maximal and therefore control of infection is difficult Patients presenting with erythema infectiosum are no longer infectious and do not require isolation Patients with aplastic crisis may be infectious at the time of presentation and should not be cared for near other atrisk patients with hematologis immunodeficiency or who are pregnant CANCER AND VIRUSES CELL CYCLE The cell cycle is an ordered set ofevents culminating in cell growth and division into two daughter cells Nondividing cells not considered to be in the cell cycle The stages are G1SG2 M The Gl stage stands for quotGAP 1quot The S stage stands for quotSynthesisquot This is the stage when DNA replication occurs The G2 stage stands forquotGAP 2quot The M stage stands for quotmitosisquot and is when nuclear chromosomes separate and cytoplasmic cytokinesis division occur Each cell cycle consists of two phases called the interphase and the M phase The interphase is further separated into three phases ofa period of DNA replication or S phase which separates two growth phases or Gl and G2 phases The duration of interphase may be very short and consists only ofthe time period necessary for DNA replication such as the case ofembryonic cells On the other hand some cells are forever locked in interphase such as differentiated cells like skeletal muscle and neurons These cells are said to be in G0 state to distinguish from the Gl phase which implies the cell will eventually undergo DNA replication The typical adult mammalian cells that undergo cell division generally have cell cycles of 1236 hrs with most ofthe variability during the Gl phase In contrast when cells divide the Mphase or the mitotic phase generally lasts 3060 minutes regardless of the cell type Cyclins are cytoplasmic proteins which stimulate transition ofa cell from Gl to S and also entry into M phase The cyclins are found complexed with specific protein kinases The complex of a cyclin and a protein kinase is called a maturation promotion factor MPF that is a dimer ofa protein kinase and cyclin Because this factor is the same as the one controlling mitosis MPF is now also called mitosis promoting factor Cdk cyclin dependent kinase adds phosphate to a protein along with cyclins are major control switches for the cell cycle causing the cell to move from Gl to S or GZ to M CANCER AND TUMOR SUPPRESSOR GENES The single most important characteristic ofa cancer cell in the body or a culture dish is the loss of growth control The rate of growth and division is not appreciably different between normal and cancer cells However normal cell growth and division are responsive to stimulatory and inhibitory influences in the environment but cancer cells often behave independent of the in uences There are a large number ofstructural and biochemical differences between normal and cancer cells but differences between cancer cells make a quottypicalquot cancer cell impossible to describe Some ofthe most striking phenotype of a cancer cell is the transformation within the chromosomes Normal cells are fastidious in maintaining their normal chromosome complement during growth and division but cancer cells often have highly aberrant numbers ofchromosome or a condition called aneuploidy The cytoskeleton of normal cells generally contain well organized arrays of microtubules micro laments and intermediate filaments but cancer cells often have reduced and disorganized arrays of cytoskeletal elements Cancer cells often express different membrane proteins which changes the adhesivity ofthe cells The loss of adhesivity is re ected in the increased motility of cancer cells Because ofthe loss of response to inhibition of growth by neighboring cells and the increased motility cancer cells in culture often overgrow each other rather than remain in a monolayer A number ofdiverse chemicals ionization radiation and DNA and RNA viruses have been shown to be able to cause cancer and referred to as carcinogens AII ofthese agents have a common property of causing changes in the genome but such changes alone are usually insuf cient for the development of cancer The transformation of cells to the cancer state usually occurs with two distinct phases of nitiation and promotion Since gene mutations are stable and inheritabIe an initial exposure to a mutagenic substance initiator may not be suf cient for transformation but a subsequent treatment promoter that stimulates proliferation may then result in tumor formation The genes that have been linked directly with carcinogenesis fall into two classes oftumor suppressor genes and oncogenes Tumor suppressor genes encode proteins that restrain cell growth and are part ofthe negative control of cell cycle regulation There are two well studied examples oftumor suppressor genes retinoblastoma Rb and p53 The retinoblastoma tumor develops in young children from neuroblasts of the developing retina Neuroblasts are embryonic neural cells undergoing rapid rounds of cell division thus its growth regulation is susceptible to any perturbation fa neuroblast escapes regulation then it would give rise to a tumor mass The gene whose Ioss thus appears to be critical for development ofthe cancer is called the retinoblastoma or Rb gene In normal healthy individuals with two good Rb genes even ifone should spontaneously inactivate no harm is done RetinobIastoma is a hereditary cancer and in individuals with an inherited mutant Rb gene the inactivation ofthe good gene copy would now lead to tumor formationThe gene product has been cloned sequenced and identified It is a protein expressed in all cells that bind transcription factors E2F regulating DNA replication and gene expression During the cell cycle Rb protein is phosphorylated by Cdk to release its binding of E2F and permit the GI to S transition and replication occurs but this regulation is lost in mutant cells that do not express this gene product Since the Rb gene product is a general cell cycle regulator one would suspect it to be associated with other cancers and indeed the loss of Rb is now associated with many different cancers A second tumor suppressor gene is p53 Individuals who inherit only one functional copy ofp53 are also predisposed to cancer The protein product has also been identi ed and it regulates the expression ofanother protein p21 a key kinase that inhibits the cdc2 kinase This prevents cells from prematurely entering S phase especially ifthe DNA has lesion Thus DNA damage will stimulates the production of p53 to arrest the progression ofthe cell cycle until the lesion is repaired fthe DNA damage is too severe the p53 protein directs the cell toward apoptosis or cell death fthe ceII Iacks functional p53 protein and the cell is able to survive the accumulated gene damage they progress toward an increasing malignancy This loss of correlation between complete DNA replication and cell division may explain the aneuploidy of many cancers CLASSES OF TUMOR VIRUSES Viruses and cancer Much ofwhat we know about the genes involved in the development of cancer is attributable to research into DNA and RNA viruses There are seven families of viruses associated with tumors 1 RNA and 6 DNA families The DNA viruses include the hepadnaviruses the polyomavirus the papillomaviruses the adenoviruses the herpesviruses and the poxviruses The RNA virus family is the retroviruses sub group oncoviruses VIRAL TRANSFORMATION The changes in the biological functions ofa cell that result from REGULATION ofthe cell s metabolism by viral genes and that confer on the infected cell certain properties characteristic of NEOPLASIATRANSFORMATION Among the many altered properties of the TRANSFORMED CELL are Loss ofgrowth control loss of contact inhibition in cultured cells Tumor formation Mobility Reduced adhesion Transformed cells frequently exhibit chromosomal aberrations ONCOGENE A gene that codes for a protein that potentially can transform a normal cell into a malignant cell An oncogene may be transmitted by a virus in which case it is known as a VIRAL ONCOGENE DNA TUMOR VIRUSES ln permissive cells these viruses produce infectious progeny lytic life cycle In cells non permissive for replication the viral DNA can often integrate into the cell chromosomes usually but not always at random sites A typical example is papovaviruses where only the early regulatory proteins such as largeT will be expressed in nonpermissive cells from a copy ofthe viral genome integrated within the cellular genome Papovaviridae Papovaviruses 1 PAPILLOMAVIRUSES Although there a re more than 50 different types of papilloma viruses not all are associated with cancers Vulvar penile and cervical cancers associated with type 16 and type 18 papilloma viruses and others but the most common genital human papilloma viruses HPV are types 6 and 11 2 POLYOMA VIRUSES Simian virus 40 SV 40 virus causes sarcomas in juvenile hamsters Polyoma virus causes leukemias in mice After integration into host DNA only EARLY FUNCTIONS are transcribed into mRNA and expressed as a protein product These are the TUMOR ANTIGENS 3 AdenoviridaeADENOVIRUSES These viruses are highly oncogenic in animals Only a portion of the virus is integrated into host genome This portion codes for early functions E1A region contains the oncogenes that code for several tumor antigens No humans cancers have been unequivocally associated with adenoviruses E1A gene product early nonstructural protein binds to the tumor suppressor protein pr while the early protein EtB binds to the tumor suppressor protein p53 Again when these viruses infect nonpermissive for lytic replication cells they can integrate part oftheir genome into cellular genomes and express early proteins such as E1A and E18 4 Herpesviridae HERPESVIRUSES EpsteinBarr virus infectious mononucleosis kissing disease This virus is associated with Burkitt39s lymphoma nasopharyngeal cancer B cell lymphomas in immune suppressed individuals such as in organ transplantation or HIV and 5Hodgkin39s lymphoma EBV can cause lymphoma in Marmosets and transform human B lymphocytes in vitro Kaposi s sarcoma associated Herpesvirus KSHV or HHV8 This virus is intimately associated with Kaposi s lesions The virus carries a number of genes that can promote tumor formation including chemokine genes and lymphokine analogs 5 Hepadnaviridae HEPATITIS B VIRUS This virus is intimately involved with liver cirrhosis Liver regeneration after destruction by the virus is thought to promote tumor formation The viral X gene which is a potent transactivator ofcellular genes is suspected to be involved in cancer formation INACTIVATION OF TUMOR SUPPRESOR PROTEINS IN ADDITION TO THEIRABILITY TO INTEGRATE INTO CELLULAR GENOMES CODE FOR VARIOUS PROTEINS THAT AFFECT CELLULAR GROWTH ETC DNA VIRUSES SPECIFICALLY TARGET AND INACTIVATE TUMOR SUPPRESSOR PROTEINS PRB AND P53 Interaction of viral proteins coded for by DNA viruses with tumor suppressor genes Regulatory proteins of SV 40IargeT adenovirusE1a E1 b and papilloma virus E6 E7 bind to tumor suppressor genes cause their proteolytic destruction and therefore inhibit their normal functions and cause cellular transformation and oncogenesis SV40Iarge T adenovirus E1A and papilloma virus E7 proteins bind the tumor suppressor protein pRB This binding releases the transcriptional factor E2F which activates the transcription of many genes and forces cells to go through additional cycles continue to proliferate SV40Iarge T adenovirus E1 B and papilloma virus E6 proteins bind the p53 tumor suppressor protein and inactivate it As a result p53 can not bind to DNA and initiate transcription of genes that stop the cell cycle andor induce apoptosis RNA TUMOR VIRUSES RETROVIRUSES 1 ONCOVIRINAE1tumorviruses and those with similar morphology First discovered was Rous sarcoma virus RSV a slow neoplasm in chickens Human tumor viruses HTLVl human Tcell lymphotropic virus Adult Tcell leukemia Sezary Tcell leukemia HTLV2 Hairy cell leukemia 2 LENTIVIRINAE HIV which causes AIDS belongs to this group It is much more closely related to some Lentivirinae than it is to HTLVI and HTLVII which are oncovirinae 3 Spumavirinae There is no evidence of pathological effects ofthese viruses ONCOGENES IN RETROVIRUSES ln retroviruses these were rst discovered as an extra gene in Rous sarcoma virus RSV This gene was called src for sarcoma src is not needed for viral replication It is an extra gene to those gagpolenv necessary for the continued reproduction of the virus Many oncogenes have been described by a number of laboratories Note that they are referred so by a three letter code eg src myo often reflecting the virus from which they were first isolated Some viruses can have more than one onc eg erbA erbB Here are a few ofthe most studied Rous sarcoma virus vsrc Simian sarcoma virus vsis Avian erythroblastosis virus verbA or verbB Kirsten murine sarcoma virus vkras Moloney murine sarcoma virus vmos M029 avian myelocytoma virus vmyc CELLS ALSO HAVE ONCOGENES The cellular homologs ofviral oncogenes are called protooncogenes or concs while viral oncogenes that originated from cellular oncogenes are called voncs Concs are not identical to their corresponding voncs After the gene was picked up by the virus it has been subject to mutation which generally has made it a more potent promoter of cellular growth CHARACTERISTICS OF CELLULAR PROTOONCOGENES Protooncogenes are typical cellular genes involved in cellular growth and cellular regulation Most concs are expressed by the cell at least on some occasions often when the cell is growing replicating and differentiating normally They are usually proteins involved in growth control Some transforming retroviruses do not have voncs An example is avian leukosis virus ALV ALV can integrate into the cell genome at many different sites but in ALVinduced tumors the virus is ALWAYS found in a similar position In all cases ofALVinduced tumors the viral genome is inserted near a cellular gene called cmyc Thus inserting the genome ofALV and other chronically transforming retroviruses next to a conc has the same effect as carrying in a vonc oncogenesis by promotor insertion Some functions of protooncgenes Control of DNA transcription found in nucleus myc 2 Signalling of hormonegrowth factor binding such as a tyrosine kinase src is a membrane bound tyr kinase 3 GTPbinding proteins GP ras Again may be involved in signal transduction from a surface receptor to the nucleus 4 Growth factors GF sis is an altered form of plateletderived growth factor B chain 5 Growth factor receptors REC erbB is a homolog ofthe epidermal growth factor receptor it is also a tyrosine kinase


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