HIV

history

• in 1981 unusual diseases recorded such as pneumonia pneumocystis and cartinii tumour

• individuals with disease had low CD4 t cells

• 1983 1st train isolated - HIV1

• 3 years later HIV2 named and noticed a pattern of virulence in diff areas —> HIV2 is less virulent and found more in Africa

• initiated from chimpanzees and sooty mangees but doesnt cause symptoms in any of them

• 1 million deaths a year

• primarily heterosexual transmission

strains and variants

• strains 1 and 2 but many diff subtypes of HIV1

• very high mutation rate which gives rise to variants

• variants differ in importance due to the cell types they infect

• all have similar structure/cell cycle

HIV structure

• part of teh lentivirus family

• its a retrovirus so has RNA genome

• 2 molecules of ssRNA each bound by a molecule of reverse transcriptase

• within the genome p10 protease and p32 integrase

• genome surrounded by nucleocapsid and containing inner layer of p24 and outer layer of p17 —> both of these together form polyprotein GAG complex

• outer proportion of virus consists of lipid from host cell memb and surface envelope proteins gp120 and gp41

genome

• GAG processed to GAG precursor —> core structural proteins p24,p18,p15

• pol region which is processed to pol precursor —> viral enzymes:

  • protease (p10)

  • reverse transcriptase p66)

  • integrase (p32)

• envelope precursor processed into gp120 and gp40 envelope structural proteins (gp120 extracellular, gp40 integral)

• also various accessory proteins such as NeR (negative regulator protein) and Vif (viral protein infectivity)

HIV GAG

• precursor which encodes the structural proteins

• processed from precursor by the protease

• membrane associated GAG associates with membrane and attracts 2 copies of virus RNA with cellular and viral protein which triggers budding of the virion

• gp120 covered in glucan side chains which helps binding of virus to host cells

replication cycle

• HIV must infect a host cell before replicating with the viral progeny leaving the cell to infect others

• HIV infects CD4 t cells —> kills them

• kills DCs and monocytes which express CD4 at lower levels

• CD4 usually aids recognition of MHC, interferes with this

• 2 stages of infection:

  • binding to host cell

  • fusion w membrane

life cycle

• binds to CD4 through gp120 and CCR5 co receptor

• nucleocapsid enters cells via fusion, unfolds and releases viral RNA

• RNA reverse transcribed to DNA

• viral DNA integrates into host genome where it lies dormant as provirus

• when cell activated viral DNA directs transcription of viral RNA

• viral proteins translated from RNA

• virus buds from the cell picking up some cell memb, complete viral particles can then infect other cells

• when the virus initially fuses with host cell, envelope proteins remain on the outside

• integrase enzyme facilitates integration of viral DNA into host DNA

• protease functions:

  • cleave the viral precursor polypeptides into individual polypeptides

  • host memb will only bud away when it has the gp120 proteins on surface of host

hijacked immune cells

• can infect DCs by binding CD4 coreceptor

• when virus leaves the host it moves through filopodea (thin projections)

  • involves rearrangement of actin to shoot out filopodea with virus attached to the end

  • allows them to attach to t cells

  • good for spreading the virus as DCs patrol the body

• however, the binding of gp120 not enough to infect cells

• originally thought that gp41 component of envelope protein binds to a 2nd protein on cell surface and 2nd protein differs between HIVs

• some variants considered that gp41 binds to beta chemokine receptor CCR5 on surface of CD4 t cells, monocytes and DCs

• variants can therefore interact with all cell types —> called M trophic

• other HIv variants bind to another chemokine receptor CXCR4 present on CD4 t cells but not DCs or monocytes —> t trophic variant

binding - new model

• gp160 is a fusion glycoprotein —> trimer formation so 3 molecules arranged together in spikes

• can overcome energy barrier to fuse membranes

• during early fusion gp160 cleaved into gp41 which remains noncovalently attached to gp120 (gp41 remains in high energy state with fusion peptide buried inwards)

• gp120 binds host cell surface to receptor and CD4 causes conf change in gp120 which allows it to bind CXCR4/CCR5 chemokine receptors

• gp41 released from high energy state and fusion peptide springs out towards memb bridging teh divide between virion and host membrane

• gp120 dissociates away and gp41 transiently becomes integral component of both membranes

variants

M TROPHIC

• can bind to DCs, macrophages, t cells

• gp120 binds CCR5 and CD4

• accessory protein MIP1 alpha aids binding to chemokine receptor

• non functional mutant allele of CCR5 (deletion of 32bp) found at high frequency in north american populations which protects against M trophic virus

• insertions in promoter region of MIP1 alpha (ligand for CCR5) found in 1 in 5 indians

• 2 alleles of CCR5 - 1 is mutated in 10% northern europeans and results in slower progression to AIDS

• approx 1% have null allele —> resistant

T TROPHIC

• binds CD4 then CXCR4

• SDF1 stromal factor aids viral binding

• point mutation (guanine for Adenine) in SDF (ligand for CXCR4 reported in 40% of healthy asians

• emergence of T trophic variants leads to rapid progression to AIDS

replication

• once inside the cell the nucleocapsid is removed and reverse transcriptase copies RNA into DNA(poor fidelity and no proofreading)

• viral DNA integrates into host cell DNA where its known as provirus (RNAse H degrades copied RNA templates)

• this stage of the viral infection known as the latent phase —> can remain dormant for a long time

• virus production is initiated by cellular TF such as NFKb which is upregulated in active T cells

• viral proteins translated using host cell protein synthesis

3 stages of infection

1. infection

2. latent stage

3. development of AIDS

1. infection

• most people show no symptoms

• about 15% develop symptoms reminiscent of flu

• some patients have swollen lymph nodes with no other symptoms

• antibodies to HIV produced in process called seroconversion

2. latency

• can last months to years

• average time to develop to AIDS is 10 years

• not everyone with HIV develops to AIDS

3. development to AIDS

• weight loss, night sweats, diarrhoea

• unusual infections such as oral thrush, herpes, shingles

• AIDS defined as appearance of major opportunistic infections or by a drop in CD4 T cell count to below 200 cells/uL of blood

• without treatment causes death due to combination of infections

  • pneumocystis carinii in Europe, USA, Oceania

  • cryptosporidium/TB more common cause of death in Asia/Africa

events associated with HIV infection

• causes profound immunosuppression

• before this IS generates powerful response to HIV

• hallmark of HIV is low CD4 but also shows in lymphnodes:

  • increased distribution of normal lymph node architecture

  • influx of CD8 T cells

  • eventual loss of germinal centres

dendritic cells

• hitches ride on DCs to lymph nodes

• but HIV exploits DCs —> picked up by mucosal DCs and avoids being killed

• the Dcs recognise siayllactose head on GM3 ganglioside (glycospingolipid) and virus is taken in

loss of CD4 function

• loss of T cell function associated with effects of the virus (separate from low T cell count)

• consequences is reduced ability to mount delayed type hypersensitivity 4 reaction (the activation and action of T cells)

• as infcetion progresses reduced ability to generate AB response

when virus is exposed to the body

• passes through epithelium where it meets and binds to DC with sialyllactose glycosphingolipid

• taken to lymph nodes where it interacts with T cells

antibody response

• increase in serum IgG despite impaired AB response

• reflects abnormalities in ability to regulate immune response

• also a sign of general immune dysfunction

• increased antibodies to:

  • RBCs

  • sperm

  • myelin

• also more allergic response

immune response

• after the initial infection viremia: rapid viral replication

• HIV IC pathogen therefore expected to initiate strong CD8 response

• theres a strong AB response to gp120 envelope and p24 protein of nucleocapsid

• also powerful CD8 t cell response against gp120, p24 and some reverse transcriptase enzymes

• immune response effective and 99% of virus is killed —> virus enters latent stage

• even during latency active viral replication (level of virmemia a good indicator for how fast will progress to AIDS)

  • cells in lymph node a lot more productively infceted than in blood

• overall T cell no declines slowly because

  • t cells destroyed but replaced daily

  • IS eliminates 30% of the viral load every day but its rapidly replaced

• all these indicate that virus is more active than initially though during latency —> clinically latent, actually dynamic

why does the immune system not clear virus

• unique features of HIV

  • fast replication rate

  • hides as a provirus

  • high mutation rate of antigen

CD4 t cell loss

• combination of direct killing and destruction of virally infected cells of immune system

• infceted CD4 cells express viral antigen (viral peptides on MHC1 or soluble gp120)

• due to expression of viral antigen can be killed in a no of ways:

  • lysis

  • antibody mediated destruction

  • antibody directed killing involving a macrophage

  • CD8 cytotoxic t cell

CD4 killing mechanism

ANTIBODY AND COMPLEMENT

• anti gp120 binds to gp120 bound to CD4 resulting in complement fixation and activation and cell lysis

ANTIBODY DEPENDENT CELL MEDIATED

• macrophages and NK cells have Fc receptors which binds to Fc region of gp120 antibody and kills cell

CD8 CYTOTOXIC T CELL

• kill virally infected cells expressing HIV antigen peptides in association with MHC 1

apoptosis vs pyroptosis

• when HIV infects permissive spleen T cell —> caspase 3 mediated apoptosis

• when HIV abortively infects CD4 t cells (fail to complete reverse transcription —> accumulate incomplete HIV transcripts detected by IFI16

• causes caspase 1 activation and caspase 1 dependent pyroptosis

• 95% of t cell death in lymph nodes is driven by pyroptosis

• pyroptosis highly inflammatory and brings more cells to be infected

pyroptosis CD4 t cell location

• progression to AIDS driven by pyroptosis mediated T cell depletion

• peripheral blood vesses have fewer HIV reverse transcriptase transcripts and so can resist pyroptosis

• when co cultured with lymphoid tissues theres increases levels of NFKb, IFI16 and reverse transcrition which leads to icreased pyroptosis of blood derived cells

• environment fo the lymphoid tissues shapes the HIV response:

  • activated lymphnode —> apoptosis

  • resting lyphnode —> abortative infection pyroptosis

  • resting RBC —> abortive infection survival

• CD4 cells killed and so its unable to generate an immune response against the virus

• rate of production of new CD4 t cells never matches the rate of destruction

• eventually the no of cells so low it causes immunosupression

therapies

• first drug is AZT reverse transcriptase which inhibits replication

• high mutation rate makes it difficult to treat

• drugs used now:

NUCLEOSIDE ANALOGUE INHIBITORS

• lack a 3’ Oh group on decarboxylase moiety so DNA synthesis stopped by chain termination

• competitive substrate inhibitors

NON NUCLEOSIDE ANALOGUE INHIBITORS

• not encorporated into viral DNA but inhibit movement of protein domains for RT

• non competitive

PROTEASE INHIBITORS

• inhibit AS of aspartic protease used to cleave viral precurusor proteins

• peptide linkage replaced by uncleavable hydroxyethene group

• so the virus cant replicate

combination chemotherapy

• raise CD4 t cell levels

• problems associated:

  • 3 drugs toxic to bone marrow/gut

  • complicated and intrusive dose of drug

  • expensive

  • successful at treatment but not clearance

WHY CAN IT NOT BE CLEARED?

• Vif accessory protein degrades IFN alpha which interferes which Jak and stat signalling pathway

• Vif degrades STAT1 and 3 from IFN alpha pathway

• no transcription of genes

new strategy

• chemical drug PA457 stops assembly of nucleocapsids prventing them from being clipped and put into position

• results in an exposed virus which is a lot easier to recognise and kill

vaccines

• initially 2 types of vaccines considered

PROPHYLLACTIC

• protective for people who dont have the virus

• issues with prophyllactic vaccines

  • too many subtypes and variants with high mutation rate

  • doesnt affect animals so no models to test the vaccine

  • chronically progressive —> would take years to see effects

THERAPUTIC VACCINES

• AIDS intially considered disease of immunosuppression where HIv causes bad immune response

• aim to boost immune repsonse

• now we know that HIV+ people ahve a string immune response its not effective

some approaches

ANTIBODIES

• using antibodies to target gp120 and block CCR5 binding

• there are several similar sequences which are conserved in survivors of AIDS —> aim to target conserved region with ABs

• however with change in epitope its difficult for AB to lock on

MICROBIOCIDES

• glycerol monolaurate blocks growth of microorganisms such as staph and chlamydia

• cells exposed to GM1 block production of moleules that appear during inflammation and that are thought to increase susceptibility of HIV infection