Retroviruses and AIDS

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Retroviruses and AIDS
Dr Amanj Saeed MB.CH.B, MSc, PhD
amanj.saeed_at_krg.org
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Discovery of retroviruses

• Retroviruses possess a unique enzyme known as RT
  (reverse transcriptase)
• RT uses viral RNA as a template for making DNA
  copy which integrate in to the chromosome of the
  host cell and serves either as basis for viral
  replication or as oncogene.
• Howard Temin and David Baltimore received Nobel
  Prize for discovery of RT enzyme.

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Discovery of HIV

• In 1981 new clinical syndrome characterized by
  profound immunodeficiency was recorded in male
  homosexual and termed AIDS.
• Unusual prevalence of Pneumocystis carinii
  pneumonia in in a group of young previously
  healthy male homosexual.

Kaposis sarcoma (rare cancer) in previously
healthy male homosexual??
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Discovery of HIV

• First isolation of HIV-1 made by Luc Montagnier
  and Barre-Sinoussi at Pasteur institute in Paris
  in 1983.
• This observation is confirmed by Robert Gallo in
  the USA.

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Discovery of HIV

• HIV-2 isolated from mildly immunosuppressed
  patient n west Africa.
• 5000 cases of HIV-1 cases per Day?
• 41 million people have been infected world wide.
• HIV-2 account for 4.5 of HIV cases.

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Retroviridae
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Lentiviruses
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Primates infected with lentiviruses
gt 30 species of African primates naturally
infected with SIV
SIV infections natural acquired not
known

• Natural infections
• gt50 of adults
• nonpathogenic
• Chimpanzee the only ape

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Primate Lentiviruses
HIV-2
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Photograph by Karl Ammann
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Photograph by Karl Ammann
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Properties of HIV

• Classification
• The family Retroviridae is named for RT.
• (Retro Backwards)
• Seven genera is now recognised (only two of them
  cause disease in human)
• Lentivirus containing HIV-1 and HIV-2,
  characterised by
• Cone shaped Nucleocapsid, absence of
  oncogenicity, and the lengthy and insidious onset
  of clinical signs.

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Properties of HIV

• BLV-HTLV retroviruses contain HTLV-I and II
  characterised by ability to cause tumours rather
  than immunosuppression.
• Spumavirus Causes characteristic foamy
  appearance in infected primate cell culture.
  (they are not pathogenic).

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Global estimates for adults and children ? 2010
People living with HIV 34.0 million 31.6 million
35.2 million New HIV infections in 2010 2.7
million 2.4 million 2.9 million Deaths due to
AIDS in 2010 1.8 million 1.6 million 1.9
million
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Adults and children estimated to be living with
HIV ? 2010
Eastern Europe Central Asia 1.5 million 1.3
million 1.7 million
Western Central Europe 840 000 770 000 930
000
North America 1.3 million 1.0 million 1.9
million
East Asia 790 000 580 000 1.1 million
Middle East North Africa 470 000 350 000 570
000
Caribbean 200 000 170 000 220 000
South South-East Asia 4.0 million 3.6
million 4.5 million
Sub-Saharan Africa 22.9 million 21.6 million
24.1 million
Latin America 1.5 million 1.2 million 1.7
million
Oceania 54 000 48 000 62 000
Total 34.0 million 31.6 million 35.2 million
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Estimated number of adults and children newly
infected with HIV ? 2010
Eastern Europe Central Asia 160 000 110 000
200 000
Western Central Europe 30 000 22 000 39 000
North America 58 000 24 000 130 000
East Asia 88 000 48 000 160 000
Middle East North Africa 59 000 40 000 73
000
Caribbean 12 000 9400 17 000
South South-East Asia 270 000 230 000 340
000
Sub-Saharan Africa 1.9 million 1.7 million 2.1
million
Latin America 100 000 73 000 140 000
Oceania 3300 2400 4200
Total 2.7 million 2.4 million 2.9 million
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Estimated adult and child deaths from AIDS ? 2010
Eastern Europe Central Asia 90 000 74 000
110 000
Western Central Europe 9900 8900 11 000
North America 20 000 16 000 27 000
East Asia 56 000 40 000 76 000
Middle East North Africa 35 000 25 000 42
000
Caribbean 9000 6900 12 000
South South-East Asia 250 000 210 000 280
000
Sub-Saharan Africa 1.2 million 1.1 million 1.4
million
Latin America 67 000 45 000 92 000
Oceania 1600 1200 2000
Total 1.8 million 1.6 million 1.9 million
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Children (lt15 years) estimated to be living with
HIV ? 2010
Eastern Europe Central Asia 17 000 14 000
23 000
Western Central Europe 1400 lt1000 1800
North America 4500 4000 5800
East Asia 16 000 11 000 21 000
Middle East North Africa 40 000 27 000 52
000
Caribbean 16 000 12 000 19 000
South South-East Asia 160 000 110 000 210
000
Sub-Saharan Africa 3.1 million 2.8 million 3.5
million
Latin America 42 000 30 000 54 000
Oceania 4600 3600 5800
Total 3.4 million 3.0 million 3.8 million
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Estimated number of children (lt15 years) newly
infected with HIV ? 2010
Eastern Europe Central Asia 2200 1700 2900
Western Central Europe lt100 lt200
North America lt100 lt200
East Asia 2100 lt1000 3800
Middle East North Africa 6800 4800 8800
Caribbean 1200 lt1000 1700
South South-East Asia 20 000 14 000 28 000
Sub-Saharan Africa 350 000 300 000 410 000
Latin America 3500 2100 5000
Oceania lt1000 lt500 lt1000
Total 390 000 340 000 450 000
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Estimated deaths in children (lt15 years) from
AIDS ? 2010
Eastern Europe Central Asia 1200 lt1000
1800
Western Central Europe lt100 lt200
North America lt100 lt200
East Asia 1100 lt1000 1700
Middle East North Africa 3900 2700 5000
Caribbean 1000 lt1000 1300
South South-East Asia 14 000 8300 20 000
Sub-Saharan Africa 230 000 200 000 260 000
Latin America 2400 1300 3500
Oceania lt500 lt500 lt500
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Morphology of HIV

• HIV particle is 100-150 nm in diameter.
• Outer envelope of lipid penetrated by 72
  glycoprotein spike (the lipid envelope protein)
• The envelope protein is composed of two subunits
  the outer glycoprotein knob (gp120) and
  transmembrane protein (gp41)
• The receptor binding site for CD4 is present on
  gp120 as well as very important antigen such as
  V3 loop.

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Morphology of HIV

• The inner surface of virus lipid envelope is
  lined by matrix protein (p17)?.
• There is also abundant cellular proteins in the
  lipid envelope (MHC class I and II) antigens.
• In HIV-1 the lipid envelope encloses an
  icosahedral shell of protein (p17), within which
  is a vase or cone shaped protein core (p24, p7,
  and p9) containing two molecules of positive
  sense ssRNA
• The RNA genome is associated with several copies
  of RT, integrase, and protease.

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HIV genome

• Positive sense ssRNA genome
• The genome is approximately 10kb in size
• The genome contain control genes which can
  enhance viral replication
• rev regulator of virus
• tat transactivation.
• vif viral infectivity
• repressor genes
• nef negative factor

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HIV genome

• The genome is flanked at each end by LTR
• 3 LTR has the polyadenylation signal and 5LTR
  has the enhancer promotor sequence for viral
  transcription.
• The pol gene code for RT, integrase and protease.

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The HIV-1 genome
rev
vif
nef
tat
5 LTR
3 LTR
vpu
vpr
pol
gag
env
p17 matrix antigen p24 capsid antigen p6/7
nucleocapsid
reverse transcriptase protease integrase
envelope glycoprotein (gp120) transmembrane glycop
rotein (gp41)
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HIV genome

• HIV binds to specific receptor on the surface of
  CD4 T lymphocytes (T-helper cells)
• It also infects
• B lymphocytes
• Macrophages
• dendritic cells
• brain cells.
• Second subsidiary receptor belongs to chemokine
  receptor family CXCR4 on the T-cells and CCR5 on
  the surface of macrophages.

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Retroviruses
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HIV lifecycle
maturation
virus binding
fusion
virion assembly and release
ssRNA ()
reverse transcription
translation of viral proteins
dsDNA
nuclear transport integration
transcription
nucleus
cytoplasm
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HIV life cycle

• After attachment the virus penetrate the cell by
  fusion from without (Mediated by gp21 and gp41)
• Synthesis of viral cDNA starts when the virion
  enters the cell cytoplasm.
• The viral RT enzyme directs the synthesis of cDNA
  strand (the minus strand) using host positive RNA
  as a primer and the viral genomic RNA as a
  template.
• Viral RNAse enzymatically remove the viral RNA
  while the RT synthesize the second DNA strand
  (plus strand).

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HIV life cycle

• Viral dsDNA will enter the Nucleolus of the host
  cell as a pre-integration complex (compose of
  viral protein M, Vpr, integrase, and dsDNA)
• the integration of dsDNA to the host chromosome
  occurs (forming pro-viral DNA)
• After integration viral and cellular factors are
  needed to activate HIV transcription.
• Initial expression of viral RNA is stimulated by
  vpr and further stimulated by cellular
  transcription factors .

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HIV life cycle

• The primary RNA transcript is spliced to give 30
  plus strand viral mRNAs.
• Viral and cellular factors are required for early
  and late viral protein expression.
• Early viral gene product include (tat, rev, and
  nef), accessory viral proteins (vif, vpr, and
  vpu)
• Late viral gene products include (gap, pol and.
  env).

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HIV life cycle

• Assembly of new virion can begin by proteolytic
  cascade by viral proteases.
• Different viral structural proteins begin to
  assemble with the p24 as a core and p7 enclosing
  viral RNA.
• Viral genome assemble in the cytoplasm.
• Retroviruses including HIV are release from the
  infected cells by budding from the infected
  cells.
• The pro viral DNA may reside quietly in the
  chromosome for years.

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Genetic Variability

• RT has NO proof-reading mechanism therefore
  mutations (point point mutations and
  deletions/insertions) occur
• Quasispecies swarm of genetically distinct yet
  related viruses

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Effects of Variability

• Immune escape by changing/masking antigenic
  determinants
• CTLs and Abs
• Resistance to anti-retroviral drugs
• Point mutations in enzymatic proteins
• RT - resistance to nucleoside and non-nucleoside
  analogues
• Protease - resistance to protease inhibitors
• Altered cytopathogenicity
• Env and particularly V3 mutations alter
  co-receptor usage
• Different cell tropism, eg. Macrophages, T-cells,
  glial cells, langerhans cells etc.
• Different tissue tropisms, e.g. brain

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Integration

• Double stranded cDNA (provirus) migrates to
  nucleus
• Can exist extra-chromosomally as linear or
  circular form
• Can integrate via the enzyme integrase

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Activation

• Once integrated the provirus responds to cellular
  nuclear factors e.g. SP1, NF-kB
• Mediated through control regions in the 5 LTR
• Once active viral factors take over
• Transactivation then control of RNA splicing
  events

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Translation

• Translation I.e.viral protein production
• Virus release via budding on cell membrane
• Morphological characteristics of budding virus is
  used for classification
• type C, Type D morphology refers to morphology of
  budding/maturing virus

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Summary

• HIV member of the Retroviridae family (reverse
  transcriptase)
• Entry mediated by CD4 plus co-receptor
• Reverse transcription leads to errors
• Virus can become integrated into chromosome (can
  be latent)
• Transcription short (spliced) then long RNAs
• New virus buds at surface
• Three main targets for therapy