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Shandia Deloach

HIV: Treatments and Immunity

July 1, 2010

Submitted in partial fulfillment of the requirements for Master's in Science

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Abstract

HIV incidence has increase steadily over the time since it was first discovered in 1981.

Progress in treatment techniques is making what was once a death sentence into more of a

chronic disease. Studying the mechanisms of infection can give key insight into new targets for

treatment. Also looking at the mechanisms that allow both long term non-progression, and

immunity can illuminate new avenues in the development of new treatments and preventative

measures. Some key molecules involved in the process include antimicrobial polypeptides, and

chemokine receptors.

As more information is revealed about the structure of the virus and it replicative

process it allows for the development of new drugs to treat the disease. One of these new

targets is the chemokine receptor CCR5. The development of antagonists to this receptor may

block HIV entry. Prevalence data has also yielded some geographical areas of anomalously low

levels of the disease. This has lead to the study of what differs among these populations and

one popular theory is that it has to do with the amount of algae consumed in these areas. Algal

extracts can become a popular arena of study especially as they appear to block viral replication

in the gut where a large amount of lymphoid tissue is located. These have the potential not

only to be inexpensive and effective drugs but also to target an area of infection previously left

untouched.

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It is commonly known that the human immunodeficiency virus (HIV) attacks the immune

system. It is this systematic destruction of the immune system that causes a person to become

immune deficient and vulnerable to diseases not seen in the immune competent (those with

functioning immune systems). In order to understand how HIV affects the immune system it is

necessary to first understand the normal function of the immune system.

h   

 

The purpose of the immune system is to provide both immediate and long-term

immunity to foreign agents such as pathogens (v9). It does this through a variety of methods,

the first of which is physical barriers to infection, which include the intact skin and mucous

membranes (7, 17, 1v9). If the pathogen manages to breach these barriers it is then the innate

immune system that attacks the invaders. The innate immune system mounts a nonspecific

response to the foreign agents. It recognizes pathogen invasion by a set of conserved pattern

recognition receptors (PRRs). PRRs work by recognizing repetitive motifs called pathogen

associated molecular patterns (PAMPS) (142) that are common to potential pathogens, as well

as markers of cellular damage called danger associated molecular patterns (DAMPs) (v9). An

example of a PAMP would be lipopolysacccharide (LPS) which is found in the outer membrane

of gram negative bacteria (v4). DAMPS are host molecules released during times of stress or

damage, such as extracellular ATP, they are distinguished from PAMPS as they are derived from

͞self͟ molecules (v9).

The innate immune system includes cellular components such as natural killer cells and

neutrophils, and molecular components such as the complement cascade and interferons (v9).

v
Antimicrobial polypeptides (AMPs) may also be considered as being a part of innate immunity

as they are also mounted as part of the nonspecific responses (121). AMPs are produced by

macrophages, neutrophils, epithelial cells, haemocytes, fat body, reproductive tract and others.

Natural killer (NK) cells mediate the lysis of infected cells through the release of the cytotoxic

granules perforin and granzyme (106). Perforin, is a 70 kDa protein which is dependent on the

presence of Ca2+, the monomers bind and insert into the target membrane where they

polymerize to form aggregates, which form a pore of between 5-20nm (88, 116). Granzyme or

granule enzyme, a serine protease, is taken up into endosomes to enter the cell. Interaction

with perforin is necessary for its release into the cytosol from the endosomes (55, 148).

Interaction of perforin with granzyme triggers apoptosis of the affected cell (116). Neutrophils

kill pathogenic microbes by engulfing and destroying them (17, 67).

The complement cascade is generated by the complement system; its two primary

functions are lysis and opsonization. The complement system relies on a series of protein

interactions in the blood to mediate its function (61). There are three pathways for the

activation of complement system, classical, mannose binding lectin (MBL), and alternative, all

converging in the activation of the membrane attack complex (MAC) that triggers cell lysis

(1v2). The MAC is a group of complement proteins that can lyse target cells, bacteria, virus

infected cells, or whole viruses, by penetrating their outer membranes (1v9). Opsonization

increases the effectiveness of the immune response by targeting cells for destruction (1v2). It

does this through the attachment of opsonins, surface bound proteins that identify foreign

material (1v9).

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Interferons (IFNs), antiviral proteins formed by virus infected cells and in response to

immune interactions with microorganisms and chemicals, can be divided into two types, Type I

and Type II (15, 45). Type I are viral IFNs and include ɲ, ɴ, ʘ, and ʏ subtypes, whereas type II

includes only IFN-ɶ (15). These proteins modulate the immune response by interfering with the

ability of the virus to infect other cells (45).

The innate immune system focuses on immediate (hours to days) immunity, however, it

is insufficient for long term human survival so one of its key functions is to activate the acquired

immune system (v9). The innate immune response is also called the inflammatory response

and some cells that promote or enhance this process are known as inflammatory mediators

(v5). For long-term control of a pathogen the body mounts an adaptive (acquired) immune

response that is specific to a particular pathogen (v9).

 
   

Antimicrobial polypeptides, especially defensins, are important in the mucosal response

to disease. Other AMPs include cathelicidins and histatins and they act against bacteria, fungi

and viruses. Many of these come into action when studying the mechanisms of immunity to

HIV. Most AMPs are < 10 kDa, have a net positive charge, and are membrane active and

hydrophobic (121). They have been identified in organisms ranging from insects and plants to

humans. Based on their structure they have been divided into five groups; V-helical, cysteine

rich, -sheet, rich in regular amino acids (AA), and those with rare AAs. While defensins are

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classified as cysteine-rich, both cathelicidins and histatins fall under the category of rich in

regular amino acids (121).

Cathelicidin LLv7, for example, comes from neutrophils and epithelial cells and can

inhibit lentiviral replication as well as being chemotactic (attracting) for neutrophils, monocytes

and T lymphocytes (1v9).

?  

Defensins are a class of antimicrobial peptides (AMP), mediators of innate host defense

that are primarily involved in membrane depolarization (12). Defensins, v-6 kDa non-

glycosylated peptides, are a potential innate HIV-1 inhibitor that have been found in mucosal

secretions and blood and are effectors that link innate and adaptive immune responses (12, 62,

12v). The production of defensins is dependent upon nod (Nucleotide-binding and

oligomerization domain) signaling (v4). There are three categories of defensins in vertebrates ɲ,

ɴ, and ɽ defensins (146). The first two of these are found in humans and are differentiated

based on the structural pairing of their cysteine residues and disulfide bridges (12, 12v). The

third category ɽ-defensins were initially isolated in rhesus macaques (14v, 146).

ɲ-Defensins are further divided and named human neutrophil peptides 1-4 (HNP-1 to

HNP-4) because they are produced by neutrophils; though immature monocyte derived

dendritic cells have also been found to produce them (12). They are found circulating in the

plasma, within certain epithelial cells and are contained in neutrophilic granules (90). Some

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studies only list the first three categories as divisions of ɲ-Defensins as HNP ʹ 4 was not isolated

until later (v0, 12v). HNPs1-v have been shown to be effective against Staphylococcus aureus,

Pseudomonas aeruginosa, Escherichia coli, and also against the fungus Cryptoccocus

neoformans as well as herpes simplex virus type 1 in vitro (51).

ɴ- defensins are produced by epithelial tissues and there are four identified in humans

HBD1-HBD4. Defensins act primarily by permeabilizing cell membranes high in anionic

phospholipids (12). ɽ-defensins are 18 residue cyclic peptides that act as entry inhibitors; they

are homologues of the ɲ-defensins expressed by humans and some other mammals (146).

Pseudogenes (relatives of known genes which are presumably no longer expressed) of

ɽ-defensin have been found encoded near the other defensin genes in humans. Their signal

sequences all contain a premature stop codon that prevents them from being transcribed, this

has also been found in bone marrow mRNA (109, 146). ɽ-defensins are also called retrocyclins

and are categorized as retrocyclins 1, 2, and v. Retrocyclin 1 has been shown to bind gp120 and

CD4 protecting cells from T- (T cell specific) and M- (macrophage specific) tropic strains of HIV-1

(146).

A study done on exposed but uninfected Kenyan sex workers found that HIV

neutralization was associated with HNPs1-v in vaginal secretions, though higher levels of these

were also associated with co-infections and higher HIV acquisition (81). This may seem a little

counter-intuitive but infection with other STIs (sexually transmitted infection) increases the

amount of these HNPs while at the same time making people more vulnerable to HIV infection.

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Other proteins that have shown anti-HIV activity are the APOBEC family of proteins.

 h

The APOBEC (apolipoprotein B [apoB] messenger RNA [mRNA] editing catalytic

polypeptide) proteins are important as some members of this family provide protection against

viruses, especially HIV. The name is derived from the fact the first member of the family to be

discovered, APOBEC1, works in the intestine, and edits apoB mRNA producing apoB48 which is

the main structural component of chylomicrons (64, 111). APOBECs are the catalytic

component of a complex to deaminate cytidines on RNA or DNA (111). This deamination

changes a single cytosine to uracil to generate a premature translational stop codon. The

family contains APOBECs 1-4, cellular cytidine deaminases, and activation induced cytosine

deaminase (AID) (100, 111). The role of APOBEC2 is uncertain but its expression is specific to

skeletal and cardiac muscle (111).

The APOBECv family is unique to mammals and includes APOBECvA (hAvA) through

APOBECvH (hAvH), it also works specifically on single stranded DNA (ssDNA) rather than RNA

(58, 111). Of the seven APOBECv genes in humans hAvG has the most potent anti HIV activity

(100). Other members of the family have also been shown to have activity against other viruses

and retroelements (transposable DNA sequences) (100). The hAvG are incorporated into the

HIV virion during assembly and catalyze cytosine deamination during (-) strand synthesis. This

ultimately results in the degradation of the viral DNA and/or lethal hypermutagenesis (100).

Unfortunately, these effects are counteracted by the viral protein vif (68).

8
 



To understand how the immune system is affected by disease it is helpful to first

consider the types of cells involved in the immune process. Clusters of differentiation (CD) are

used to delineate between different types of immune cells. These are cell surface phenotype

antigen differences, of which there are over v50 (http://www.hcdm.org/) (2v). Within a

cluster, antibodies give identical cellular reaction patterns and identify the same molecular

species (74). Key in the discussion of HIV are the CD4+ T cells as the CD4 molecule is used as a

receptor for viral entry into cells. Further detail on immune cell function can be seen in Table 1.


     
Macrophage/Monocyte Phagocytosis, antigen presentation, secrete
interleukin-1 and interferons
Neutrophil Phagocytosis
Basophils Inflammatory mediator, release histamine and
heparin
Mast cells Inflammatory mediator
Eosinophils destroy antigen antibody immune complexes
Dendritic Follicular Sense changes in Bind opsonized virus
cell homeostasis and and trap it
Plasmocystoid target naïve T-cells Produce interferon ɲ
Natural Killer cell Induce apoptosis
T cells CD 4 helper secretion of interleukin-2, ɶ-IFN, and tumor
necrosis factor, proliferation of T and B cells
CD 8 cytotoxic Include suppressor T cells that reduce immune
function, and cytotoxic cells that release
cytokines to attract macrophages
Memory Found in lymphoid tissue, recognize invading
antigen from prior exposure
B cells Mature into plasma cells which produce
antibodies

   


(1v, 17, v9)

 

  

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The acquired immune response is activated as follows. First an immunogen (a

substance or organism that triggers an immune response) is captured (endocytosed) by an

antigen presenting cell (APC), generally a dendritic cell. APCs are differentiated into

͚professional͛ and ͚amateur͛ based on their ability to generate ͞signal 2͟ which involves the

expression of surface protein co-stimulatory molecules such as CD80 and CD86 that generate

this response (67, 99). Dendritic cells are considered professional due to their ability to elicit

this second signal and thus are the preferred APC (99). Dendritic cells respond to changes in

homeostasis by maturing and limiting surrounding cell activation and targeting naïve T-cells.

Activation of dendritic cells regulates antigen phagocytosis and presentation, also called ͞signal

1͟ in T cell activation (v9). The antigen, for example a viral protein, is cleaved into small

peptide fragments (representing specific viral epitopes, fragments of the antigen that will

generate an immune response) inside the APC. The APC then migrates to the lymphoid tissue

for antigen presentation (v5, 99). This epitope is then presented to B and T cells in association

with a cell surface protein called the major histocompatibility complex (MHC) (2v). MHC

molecules can be subdivided into classes I and II. B and T cells possess surface receptors that

respond to a specific epitope. In the case of T helper cells the binding of the MHC and epitope

along with the release of the cytokine interleukin 1 (IL-1) from the macrophage triggers the

activation of the cell. This activated T helper cell can then, with the aid of IL-4 and IL-5 which

are secreted by the activated T cell trigger the activation of B cells (v5). The T cell can then

mature and become a memory T helper cell (Table 1) or with the aid of interleukin 2 (IL-2) it can

trigger the activation of cytotoxic T cells. These cytotoxic T cells release cytotoxins, perforin

and granzyme that result in the death of virus-infected cells. They can also mature to become

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memory cytotoxic cells. The activated B cells react with virus immunogen on the APC and

differentiate to become plasma cells that secrete antibodies. Viruses have many methods of

evading the immune response some of which will be discussed later in regards to HIV.


 Function

!" activate T lymphocytes by enhancing the production of IL-2 and the expression of IL-2 receptors

!" generation and maintenance of T regulatory cells, activate NK cells and promote
the proliferation of B and T lymphocytes
!"# contribute to the activity of eosinophils in allergic inflammation through their capacities to
prolong eosinophil survival and to generate activated eosinophils, supports the growth of
precursors for a variety of hematopoietic cells

!"$ Induce antiparasitic and allergic immune responses, negative regulator of TH17, IL-19, IL-25, IL-
vv

!"% stimulating eosinophil production and release from the bone marrow, is chemotactic for
eosinophils and activates mature eosinophils, inducing eosinophil secretion and enhanced
cytotoxicity

!"& B lymphocytes differentiation into mature plasma cells and secrete immunoglobulins mediates
T-cell activation, growth, and differentiation, production of acute phase proteins

  




    (28)

B cells are differentiated from hematopoietic stem cells in the bone marrow and are

defined by the production of immunoglobulin (Ig) antibodies. There are five classes of

immunoglobulin, IgA, IgC, IgD, IgE, and IgG. Naïve B cells express IgM and IgD on their cell

surfaces (2v). Stimulation by T cells triggers class switching which is characterized by the

production of IgG, IgA, or IgE antibodies (2v). Memory cells are formed in the late stages of

immune activation and require a smaller threshold for activation than non-memory cells. They

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are key in providing a swift response upon re-exposure to a pathogen and are important in the

mechanism by which vaccines work (40).

Antibody Function
IgA Provides local protection to mucous membranes;
present in tears, saliva, mucus, breast milk and other
mucosal secretions
lgD Activates B-cells; assists in differentiation of B-cells
IgE Involved in allergic reactions; defense against parasitic
infections
IgG Protects against bacteria and viruses; enhances
phagocytosis and triggers complement system; can cross
placenta from mother to fetus
IgM First antibodies secreted; cause lysis of pathogens
#  

  


(17)

' 

Viruses are biological entities that infect other cells and replicate. There are an

estimated 10v1 viruses on earth (16). When outside of cells they exist as free particles and their

general structure typically includes a RNA or DNA genome surrounded by a protein shell (16).

The majority of viruses are phages that attack bacteria (16). When a virus infects a eukaryotic

cell it generally triggers apoptosis of the infected cell by either directly or indirectly activating

cellular sensors that initiate cell death (42). Host mediated lysis is one of the mechanisms by

which viruses subvert the immune system, after replicating within the cell they use this lysis as

a means to spread to other cells (41).

Viruses are classified into two major groups based on the type of nucleic acid they

contain, riboviruses (RNA-containing) and deoxyriboviruses (DNA-containing) (119). They can

12
further be divided by their genome type; double-stranded (ds) DNA, single-stranded (ss) DNA,

dsRNA, negative sense ssRNA, positive sense ssRNA and reverse transcribing (96, 119). The four

principle taxa (related clusters) of viruses as recognized by the international committee on the

nomenclature of viruses (ICTV http://www.ictvonline.org/) are Species, Genus, Family and

Order (96). Family and order are not always used as there is not always the needed relatedness

between groups. These taxa can all be classified underneath their genome type. Of particular

interest are the reverse transcribing viruses or retroviruses.

(  

Retroviruses are a family of encapsulated (within a viral envelope) RNA viruses (75).
They are about 90 -120 nm in size and consist of a two copies of the

)    ) 


* !(+ )
     
     ,
ð ð+)   ) ,+ 
,+  ) -
(2)

ssRNA genome and number of enzyme molecules required for stable infection (89). One of

these is the reverse transcriptase, encoded by the pol gene, which is an RNA-directed DNA

polymerase that is essential for viral replication. All retroviruses contain three major coding

domains which are the gag, pol, and env genes. Gag, named for group specific antigen, directs

the synthesis of proteins that form the matrix, capsid, and nucleoprotein structures, pol

contains the information for the reverse transcriptase and integrase enzymes, and env codes

for the viral envelope proteins (26, 15v). There is also a smaller coding domain pro which

1v
encodes for the viral protease. The long terminal repeat (LTR) is also a part of the retroviral

genome and it contains the promoter and enhancer regions (75). Retroviruses can be classified

as simple or complex depending on whether they carry only this information or whether they

code for additional regulatory proteins as is the case with HIV (89).

)     ) ð ð-+


.    
,+    
  
   
., +       (       /
, &  
 
   0'  ,       )ð ð


The Retroviridae family is further divided into three subfamilies containing seven

genera. Retroviruses are found in a number of hosts, as determined by its envelope

glycoprotein which controls viral tropism and promotes the membrane fusion process (89, 107).

These genera are differentiated based on the shape and location of the inner protein core (26).

These are the oncovirinae; alpharetrovirus, betaretrovirus, gammaretrovirus, deltaretrovirus,

and epsilonretrovirus, the spumavirinae (foamy) spumavirus, and lentivirinae (slow) lentivirus

(120). The first five of these are retroviruses with oncogenic (cancer causing) potential, hence

the subfamily name oncovirinae, and are all simple retroviruses containing only the gag, pol,

and env genes (26). Oncogenic viruses are considered to be directly linked to cancer (155).

These seem to be associated with the incorporation of viral oncogenes into host DNA,

modification of viral oncogenes after integration, and the modification of host genes such as

human endogenous retroviruses (HERVs) integrated into viral genomes that then act as

14
oncogenes (155). All of these are pathogenic either in their natural host or in incidental hosts

(86). The lentivirus, meaning slowly developing, family is especially relevant as the virus HIV

belongs to this family.

!   

 Lentiviruses are found in felines, ungulates (sheep, cattle, etc.), avians, rodents, humans

and primates (79) (75). They cause disease primarily by killing or inducing the loss of function

of specific cells and tissues (26). Diseases included in this family encompass HIV, simian

immunodeficiency virus (SIV), feline immunodeficiency virus (FIV) and others (79). HIV is the

only one known to infect humans (72). These viruses are characterized as being nononcogenic,

immune cell infecting, remaining permanently associated with cells, causing slowly progressive,

chronic diseases, and the replication is generally species-specific. Common features in host

interaction with these viruses are long periods of clinical latency, only a small neutralizing (virus

specific) antibody response, extensive genetic mutation and drift, neuropathology and viral

infection of some bone marrow derived cells resulting in lysis (79).

1984 Prevalence Data


Ë  È of reported cases
Men who have sex with men 71.4
IV drug users 16.9
Haitian Immigrants 5.v
Hemophiliacs 0.7
Other/None 2.7
Unknown v
$(   0' 
(27)

15
0
 

 ' 10'2

0

Human immunodeficiency virus, HIV, is a retrovirus belonging to the lentivirus family

(75). It is responsible for progressive immune dysfunction that leads to acquired

immunodeficiency syndrome, AIDS (4v, 124). The symptoms of HIV infection may include;

swollen lymph nodes, fever, chills and night sweats, diarrhea, weight loss, coughing and

shortness of breath, persistent tiredness, skin sores, blurred vision and headaches, as wells as

the development of other infections (92, 1v1). It is spread through the transfer of bodily fluids

such as; blood (92), semen (57), vaginal secretions (1v1), and breast milk (10). Only in about 1-

5È of patients does saliva contain infectious HIV and tears contain only negligible amounts of

the virus (9, 108). The virus can survive outside the body for several days to weeks depending

on the environment, initial viral titer, and growth medium. Factors such as temperature,

humidity and sunlight affect the life of the virus outside the body (1v7) influencing survival

include. A person is generally considered to have progressed to AIDS if they have specific

clinical conditions indicative of severely impaired immune function, or a CD4 cell count of less

than 200/mmv (84). In the absence of treatment the onset of AIDS generally develops 8-10

years after infection (71).

HIV was first drawn to the attention of physicians in 1981 when the Centers for Disease

Control, (CDC http://www.cdc.gov/), reported five cases of Pneumocystis carinii pneumonia in

young homosexual men in Los Angeles (57, 102). Within a year the term AIDS was coined by

the CDC to describe the combination of opportunistic infections and tumors occurring in people

with markedly reduced helper (CD4+)T-cell count (table 1) (124). Initially the disease seemed to
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be confined to a few high risk groups of people, (see table 4), however, in 198v a retrovirus

later named HIV was isolated from a patient with AIDS thus confirming suspicions that AIDS had

an infectious cause (102, 124). Finding this retrovirus led to the development of a test for HIV

infection. The immunoglobulin-G antibody test utilizes an enzyme-linked immunosorbent assay

(ELISA) to search for IgG antibodies specific to HIV in a patient͛s serum, which also allows for

the screening of donated blood for large scale epidemiological studies and for prevention of

infection from receipt of donated blood (115). Viral load is measured by assays such as the

Amplicor HIV-1 Monitor test (Roche Molecular Systems) which measures viral RNA by nucleic

acid amplification (69). There has been dissent over the naming of HIV as to the etiological

cause of AIDS, with some proposing that AIDS is caused not by HIV but rather by drugs and

other non-contagious risk factors, however, sufficient evidence to support alternate theories

has not been established (v6, 46).

The earliest documented case of HIV infection was identified from a stored plasma

sample from 1959 in the city now known as Kinshasa in the Democratic Republic of Congo

(154). The origin of the virus in humans is postulated to be the inadvertent introduction of

simian immunodeficiency virus (SIV) from a closely related primate species. In 1986 a second

strain of retrovirus called HIV-2 was isolated from patients with AIDS in West Africa (25, 124).

This strain shows a 40-60È homology at the RNA level with HIV-1 but differs significantly at the

protein level (25). Studies of HIV-2 show that it has a very close relationship with the SIV strain

that infects the primate known as the sooty mangabey (Cercocebus atys) in the same parts of

West Africa (77, 124). It is now thought that there have been at least three separate entries of

SIV into the human population. HIV-1 seems to have a greater degree of similarity with

17
chimpanzee (Pan troglodytes) strains of SIV in species whose habitat overlaps that of the

location where the HIV-1 is believed to have originated (124). Exactly how humans came to be

infected is yet to be established.

There are at least four different African primate species with their own

immunodeficiency virus variants. These include the Sooty mangabeys (Cercocebus torquatus

atys) SIVsm, African green monkeys (Cercopithecus aethiops) SIVagm, Sykes͛ monkeys

(Cercopithecus mitis) SIVsyk, and Mandrills (Mandrillus sphinx) SIVmnd (114). Also there have

been reported cases in chimpanzees (Pan troglodytes) SIVcpz and red-capped mangabeys

(Cercocebus torquatus torquatus) SIVrcm though it is unclear whether they are natural hosts

(114). In no case do these viruses appear to be pathogenic in these other primates and studies

of the mechanism behind this could give insight into dealing with HIV infection in humans (114).

It is important to note that this is not a function of viral load as there can be high viral loads in

these animals without progression of the disease to AIDS (9v). One study indicated that the

disease will eventually progress to classic AIDS if followed over a long enough period of time

(85). Despite this it is still apparent that non-human primates have some defense mechanism

that humans lack.

The main immunologic feature of advanced stages of HIV is a decrease in the number of

circulating CD4+ T-cells, which are helper T cells. This feature quickly became the main

surrogate marker for HIV-related immunodeficiency. The normal CD4+ T-cell count is >500 T

cells per mmv, v50-500 T cells per mmv is slightly depressed, 200-v50 T cells per mmv is

moderately depressed and <200 T cells per mmv demonstrates severely compromised immune

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function (97). Once absolute CD4+ T-cell count drops below 200 T cells per mmv in the

peripheral blood there is a susceptibility to characteristic, AIDS defining, opportunistic

infections. The tendency of the virus to infect CD4+ T-cells as well as other CD4 expressing cells

was explained in 1984 when the CD4 molecule was shown to be a high affinity receptor for the

virus (v1).

The time course of HIV infection can be tracked by a combination of CD4+ T cell count

and viral load. In the first weeks after initial infection there is a drop in cell count and a rise in

viral load (amount of viral RNA detected) (128). This returns back to normal or near normal over

the course of the next weeks or months. The virus then enters a period of what is generally

considered to be latency, when viral load and cell count reach a stable set point (17). This set

point differs from person to person but looking at this can give an indication of how the disease

will progress. The set point for plasma viral load can range from as few as 100 copies/mL to as

many as 10,000 copies/mL. Patients on the higher end are at higher risk of disease progression

(17). During this time viral load and cell count remain relatively steady, and this can last for

years (17). Viral shedding can occur at any time during the course of the infection but other

STIs, as well as higher viral loads lead to increased shedding (128).

) #0'" ) 


 ) 
(2)

19
     

The HIV virus is composed of a central core of cylindrical RNA which is surrounded by a

spherical bilayer membrane envelope with glycoprotein (gp) surface markers (84). Cell surface

markers enable the identification of specific cell types. The viral genome contains ~10,000

nucleotides and encodes nine genes. These include the three common to all retroviruses gag,

pol, and env, as well as tat, rev, vpu, nef, vpr, vif whose functions will be explained later. In HIV-

2 the vpr gene is replaced with the gene vpx (10v). HIV-1 entry into the cell requires interaction

with two separate membrane protein sites on target cells. These sites are the CD4+ receptor

and a 7 transmembrane (7-TM) chemokine receptor (84). This 7-TM chemokine receptor is

generally the CC-chemokine receptor 5 CCR5 or the CXC chemokine receptor 4 CXCR4 for

primary HIV-1 in vivo (127). These are named for the structure of the chemokines which they

normally bind. CC chemokines are a group of about 28 chemokines, which characteristically

have two adjacent cysteines (CC), while CXC have a variable amino acid between the two

cysteines (v2). HIV is classified as M-tropic, macrophage specific or T-tropic, T cell specific

dependent upon the type of cells it attacks. Viral tropism can change over the course of the

disease, initially infecting as M-tropic but changing to T-tropic as the virus progresses (84, 117).

CCR5 is primarily used by the M-tropic, viral strain whereas CXCR4 is more prevalent with the T-

tropic viral strain. This preferential targeting of the viral strains also gives the tropisms another

name: M-tropic is also designated R5 for use of the chemokine receptor CCR5 and T-tropic is

also designated X4 for use of the chemokine receptor CXCR4. The genes for chemokine

receptors are constitutively expressed allowing leukocytes to respond immediately to

emergencies as signaled by on-site chemokine production (110). Specifically CCR5 aids memory

20
T cells by stimulating the up-regulation of adhesion receptors and allowing migration to

inflammatory sites. This function is also carried out by CCR2 and CCRv (40). CXCR4 is

chemotactic for T cells, and is involved in B cell development in the bone marrow (v2). It also

plays a role in vascularization of the gastrointestinal tract and formation of cerebellar tissue, as

defects have been seen in these areas when it is knocked out (v2).

There are two primary strains of HIV that are the focus of most studies, HIV-1, and HIV-

2. As mentioned earlier each of these is closely related to a specific SIV strain from primates.

The worldwide pandemic of HIV is primarily due to HIV-1 so this is the primary focus (129). HIV-

1 is further divided into three groups main (M) (not to be confused with the M-tropic (R5)

designation above), outlier (O), and non-M, non-O (N). The M group causes the majority of

infections and is divided into subtypes or clades given the letters A ʹ F (52, 129). These are

differentiated by their phylogenetic ancestry or the point at which they diverged. Variance in

subtype also denotes a variance in antigenic structure, transmissibility and infectivity (129).

Half of the viral infections worldwide are C type (1).

21
) $'     

(80)

As is shown in figure v there are several steps in viral replication: binding, fusion and

uncoating, reverse transcription, integration, transcription, translation, virion assembly, and

budding and maturation. Each of these steps involves particular proteins encoded by the viral

genome in order to proceed. The chemokine and CD4 receptors mentioned earlier and other

host proteins are also involved in this process. The following is a brief overview of the

replication process. Greater detail into the mechanics of this will be given as the proteins

involved are discussed later.

In general the process of viral replication is as follows: HIV binds to the host cell

through the interaction of gp120 and CD4+ receptor site (17). Following this, interactions

between the virus and chemokine receptors trigger irreversible conformational changes (128).

After binding, gp41 facilitates the fusion of the viral envelope with the host cell membrane

22
through pore formation. Once fused the HIV viral core then uncoats releasing two single

strands of viral RNA into the cytoplasm (17). Reverse transcriptase then utilizes this ssRNA as a

template to make dsDNA. This dsDNA moves into the host nucleus where integrase promotes

its insertion into the transcriptionally active domains of the host cell DNA making proviral DNA

(17, 128). When the host cell is activated transcription of this proviral DNA into mRNA occurs.

Then mRNA exits the nucleus and aids in the production of viral polyproteins that will become

the HIV core and envelope. The enzyme protease modifies some of these polyproteins into

active proteins such as the envelope proteins gp120 and gp 41 (17). These are then assembled

into an HIV virion and then budding occurs to release HIV from the cell. Cleavage of the

budding virion is accomplished by the viral protease. It is this cleavage that marks the

maturation of the virion (128).

Ë 34$

Entry of HIV into the cells is mediated by the envelope (env) gene coding for

glycoprotein gp120 (outer envelope) and gp41 (transmembrane) molecules, which are derived

from cleavage of gp 160 and are responsible for viral attachment to the target cell (92, 150). Gp

120 surface subunits are responsible for binding to cellular receptors and gp41 acts as an

anchor (v7). The first step in this entry is the binding of gp120 to CD4 followed by binding to a

receptor on the T-cell surface that triggers conformational changes in Env; this creates or

exposes a binding site for the 7TM chemokine receptor (v8). The identity of the receptor and

the specific mechanisms behind this viral entry are not clearly understood (29).

2v
 
  

Chemokines are a varied group containing 40 ʹ 50 small molecular weight

polypeptides. Their general function involved the chemical attraction of various cells to specific

tissues (v2). They are traditionally classified according to the amino acid motif that occurs in

the first two N- terminal cysteines (v2). For the largest of these groups the amino acid motif

consists of consecutive cysteines CC and cysteines interrupted by an intervening amino acid

CXC. The CC or beta chemokines act on lymphocytes, monocytes, basophils, and eosinophils

and the CXC or alpha chemokines act mainly on lymphocytes and neutrophils (v2). The

receptors for these are in the family of seven transmembrane G ʹ protein coupled receptors

GCPRs which follow after the nomenclature of the chemokines that they respond to (v2).

These receptors range from vv9 to v7v amino acids in length (110). Their general functions

include cell migration, transcriptional activation, cell growth and differentiation (149).

As mentioned earlier viral tropism determines the type of cells that it infects and CCR5

and CXCR7 are the most common co-receptors for HIV. Studies done in vitro have

demonstrated that other chemokine receptors such as CCR2, CCRv, and CCR8 may also be used

as co-receptors for HIV (4). The M ʹtropic strain of HIV is thought to be critical in infection

establishment and maintenance and is the primary transmitting form of the virus (v2, 95).

CCR5, the primary co-receptor for the M-tropic (R5) strain, was first identified as the receptor

for CCLv, CCL4, and CCL5 (v2). It is expressed on memory T cells and IL-2 cultured T cells as well

as immature dendritic cells. The expression of CCR5 on dendritic cells is dramatically reduced

24
after lipopolysacccharide induced maturation of these cells (v2). Its normal function is to

promote chemotaxis of leukocytes towards the sites of inflammation (95).

CXCR4 is also known as Fusin, LESTR, and HUMSTR (v2). Expression of this receptor has

been demonstrated on T cells, B cells, and monocytes and mature dendritic cells, bone marrow

B-cell progenitors and in the embryonic brain (v2). The natural ligand for this receptor is

CXCL12 (v2). Exposure of resting T cells to IL-4 has been reported to increase expression of this

receptor. In mice, knockouts of the gene for this receptor or its ligand result in lethal mutations

and in abnormalities in gastrointestinal tract vascularization (v2). HIV strains using CXCR4 are

rarely transmitted but evolve as the disease progressed and are associated with poor outcomes

(105).

-  

A v2 base pair deletion in the gene that codes for the CCR5 receptor results in a loss of

expression of functional receptors (v2). This defect is present in about 1È of the Caucasian

population (v2). People homozygous for the CCR5 ɷ v2 mutation have a high degree of

immune protection from HIV due to the lack of expression of any functional CCR5 on their cell

surfaces whereas those who are heterozygous have partial protection (4, 50). This partial

protection is conveyed in an apparent delayed disease progression (22). CCR5 also seems to

play a role in the lack of pathogenicity of the SIV infection; sooty mangabeys down-regulate the

expression of CCR5 on their CD4+ T cells (50). In red caped mangabeys there is a similar

deletion in the gene coding for CCR5 and this 24 bp deletion is present in 98È of all red-caped

mangabeys (50).

25
Other mutations in the chemokine receptors that have been studied are the CCR2-64I

point mutation and the SDF1-v͛A mutation, a single nucleotide polymorphism, in the SDF1 gene

coding for the chemokine ligand for CXCR4. These mutations confer favorable prognosis to

HAART treatment and protection against the development of AIDS respectively (4). An

additional mutation in chemokine receptors is the CCR5-P1 mutation that plays a role in slowing

the rate of HIV progression (50).

-.  

Matrix protein (MA) is one of the karyophillic (attracted to the nucleus) elements

present in the viral proteome, and it is important in the nuclear import of HIV. MA, one of the

first viral proteins of HIV-1 to be related to nuclear import, utilizes nuclear localization signals

(NLS) to mediate its action (122). NLS target molecules to the nucleus of the cell, commonly

using a short stretch of basic amino acids that give an overall net positive charge (20). The NLS

connects the preintegration complex (PIC) with karyopherin ʹ ɲ (importin ɲ), and cellular

import machinery and this drives import into nucleus (56). Karyopherin ʹ ɲ is a host protein

that is part of the nuclear import machinery (122). The PIC is a nucleoprotein complex that

contains viral proteins (reverse transcriptase, MA, NC, and Vpr) and cellular factors that are

essential for the efficient integration of retroviral cDNA (18).

26
(   

Reverse transcriptase (RT) is one of three proteins derived from the pol protein from

the pol gene in retroviruses (92). It is an RNA- and DNA-dependent DNA polymerase and also

has ribonuclease H (RNase H) activity (54). Its function is to make a dsDNA copy of the viral

RNA (75). RNAse H activity triggers the degradation of the RNA genome (126). This

transcription takes place in the cytoplasm of the host cell (Figure v). The RT enzyme lacks a

proof-reading mechanism which, along with its rapid replication rate, makes the virus easily

mutated (124). This is one of the key targets for antiretroviral therapy.

 )

Integrase (IN) is a v2 kDa enzyme necessary for the integration of viral DNA into the host

cell chromosome (11). It is part of a large nucleoprotein complex, the PIC, which includes viral

DNA, Vpr, the integrase interactor protein, and several cellular proteins, such as cellular lens

epithelium derived growth factor (11, 140). IN binds to a recognition sequence found at the

ends of the viral LTRs. It catalyzes the integration of viral DNA through a v͛end processing and

strand transfer (11, 140). The third step in integration, gap repair, is mediated by cellular

proteins (11). Inhibitors have been developed that interfere with the strand transfer process

which could be beneficial in treatment of HIV (11). The lack of a functional, naturally occurring

equivalent of the enzyme against integrase makes it less likely for toxicity to develop (11).

Transactivator of transcription (Tat) and Rev are additional viral cofactors that are

essential for replication. Tat promotes elongation of the viral genome by binding to HIV͛s

transactivation response element (TAR) (6). TAR recruits positive transcription elongation

27
factor b (P-TEFb) which is necessary for HIV transcription as well as the transcription of other

cellular genes (6). Rev is involved with the nuclear export of unspliced viral RNA. Absence of

tat and cellular activation signals in resting cells is responsible for viral latency (5v). There is

currently no way to detect the difference between uninfected cells and latently infected cells.

' 

Viral protein R (vpr) encoded by the vpr gene is a small (96 amino acid) nuclear protein

that is expressed late in the virus replication cycle. It is responsible for the nuclear import of

the preintegration complex (PIC), apoptosis of cells, and aids in the reverse transcription

process, G2 cell cycle arrest and transactivation of the LTR and/or target genes. Its actions in

the transactivation process control the gene expression of HIV (140). When vpr is deleted in

vitro the virus replicates effectively in cell lines and activated CD4+ T cells but not as well in

macrophages and monocytoid (resembling monocytes) dendritic cells (58). Mutations in the

gene have been associated with long-term non-progression resulting from a decrease in

proapoptotic activity. The protein also has neurotoxic effects and defects in the gene have

been implicated in HIV-1 dementia (140).

' 

The gene vpu codes for viral protein U (vpu). It has two primary functions, degradation

of the CD4 molecule in the endoplasmic reticulum and is involved with virion release from cells

28
(Figure v) (11v). This role in virion release is cell type specific. It works by countering an

interferon-induced restriction factor, called B cell stromal factor 2 (BST-2) or tetherin, that

would normally prevent retrovirus release. Theories as to how it does this include:

proteosomal degradation of tetherin, and acting in a manner similar to nef and down regulating

the expression of tetherin (91). As tetherin works to inhibit virion release it is being looked at

as a new target of treatment.

A few of the proteins encoded by the viral genome are involved specifically in immune

evasion. Immune evasion is the process by which viruses ͚trick͛ the immune system into

helping them spread or prevent host mechanisms from destroying the virus.

'

Viral infectivity factor (vif) is a basic, 2v kDa protein that works to counteract the effect

of APOBEC by decreasing the intracellular concentration of hAvG and hAvF, preventing their

incorporation into HIV virion (100). Vif is essential for HIV propagation in vivo. It is only in the

absence of vif that the APOBEC proteins are allowed to function. Since vif plays a critical role in

eliminating an intrinsic defense mechanism it could be an attractive target for future therapy.

Vif functions by utilizing the ubiquitin-proteasome pathway to degrade APOBEC (68, 91).

Ubiquitination is a general cellular mechanism for marking damaged proteins for destruction by

the proteasome by covalently attaching a small protein called ubiquitin. This ligation is

catalyzed by a series of complex reactions utilizing ubiquitin activating (E1), conjugating (E2),

29
and ligating (Ev) enzymes (68). Ubiquitin is activated forming an E1 thiol-ester ubiquitin

intermediate. E2 catalyze the attachment to target proteins and then Ev binds and ligates that

target protein. The HIV Vif protein hijacks an Ev enzyme, specifically the Cul5-Ev inducing

APOBEC degradation (87, 100). It is a fundamental viral countermeasure to host restriction

(91).



The nef gene, one of the six small genes specific to HIV encodes the protein nef. Nef

(negative factor) has many varied functions which makes it somewhat of a misnomer for it to

be named negative factor. In vivo it is required for efficient viral replication and pathogenicity

(1vv). In vitro it affects cellular signal transduction pathways and down regulates expression of

both surface CD4 (the HIV receptor), and MHC I and II molecules (24, 1vv). Defects in the nef

and/or long terminal repeat (LTR) region (figure2) have been associated with the absence of

disease progression (140). Nef also binds and mediates the redistribution, and removal from

the cell surface, of the immune co-stimulatory molecules CD80 and CD86. These molecules in

the B7 family act as the second signal in T cell activation, triggering CD28, an immune co-

stimulatory molecule, which is expressed on naïve T cells (67). The B7 family is a family of

immunoglobulins that includes B7-1 (CD-80), B7-2 (CD-86), B7-Hv, PD-L1, PD-L2, and GL50.

They are primarily expressed on activated B cells, and macrophages (1v4), and their functions

include T cell priming, IL-2, IL-4, IFNɶ production, and inhibition of T cell proliferation and

response (8v).

v0

 

The use of antiretroviral drugs for the treatment of HIV was introduced in the mid

1980͛s (14). Treatment typically consisted of a nucleoside reverse transcriptase inhibitor, NRTI,

monotherapy (single drug). Monotherapy did not adequately suppress the virus which led to

the emergence of drug resistant HIV variants (49). The goal of antiretroviral treatment is to

reduce mortality and morbidity rates (128). Soon trials were done that experimented with

multiple drug therapy. These studies combined two NRTIs and showed a reduction in mortality

rates and progression to AIDS (14). With the advent of new classes of antiretroviral drugs such

as protease inhibitors and non-nucleoside reverse transcriptase inhibitors new multiple drug

treatments could be used. Combination therapies reduce the incidence of drug resistance and

toxicity. These multiple drug therapies are known as ͚highly active antiretroviral therapy͛, or

HAART (14).

Current drug options include nucleoside and non-nucleoside reverse transcriptase

inhibitors (NRTIs and NNRTIs), CCR5 receptor antagonists, protease inhibitors, fusion inhibitors,

and integrase inhibitors (14, v7). HAART generally includes a combination of two different

reverse transcriptase inhibitors and a protease inhibitor (1v5). Research into new

antiretrovirals is important as difficulty in maintaining adherence, drug resistance and

short/long term side effects are problems with the current drugs. Fusion inhibitors are

generally reserved for patients who have been treated for long periods of time (11). They have

been approved along with other drugs to be used in patients who despite treatment still have

residual viremia (48). Figure three shows where these drugs act in disrupting the infectivity of

v1
HIV. Though NRTIs and NNRTIs both act on reverse transcriptase they differ in their binding

sites, NNRTIs interact with the non-substrate binding site and NRTIs interact with the substrate

reverse transcriptase binding site (vv). Binding in this non-substrate site seems to lock the

reverse transcriptase into an inactive conformation. These two sites are functionally and

spatially associated and thus using a combination therapy of the two RTIs is more effective (vv).

While these therapies are good at decreasing the viral load, they have little effect on

renewing CD4+ T cell levels or immune reconstitution. This is why there is also the need for

adjuvant therapies. Adjuvant therapies work to complement the work that the antiretrovirals

do, by directly targeting the immune system (94). The most promising of these is interleukin-2

(IL-2) immunotherapy. Studies show that in conjunction with HAART there is considerable

advantage in driving a sustainable rise in CD4+ T cells (7v).

v2
 

 
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times the patient has to switch treatment regimens. In 2002 the average drug cost for one to

six switches was ~$5,978/yr or ~$498/mth and for seven to thirteen switches it was ~$7,412/yr

or ~$618/mth (101). Decisions on changing treatment are based not only on T cell counts and

viral load, but also on whether the patient is experiencing side effects from the therapy (44).

These costs have risen as new drugs are developed and treatment regimens expand beyond the

use of three drugs.

  
 

As more information is learned about the structures involved in viral entry and

replication new drugs can be made to target them. Some of the new avenues for treatment

include CCR5 receptor antagonists, stem cell transplant from a person with the CCR5 ɷv2

mutation, fusion inhibitors, and algal extracts. It has been speculated, based on prevalence

data and in vitro and limited in vivo evidence of the anti-HIV activity of algae, that algae

consumption is related to lower rates of HIV (1v6). HIV/AIDS prevalence in eastern Asia is

about 0.1È in adults, (Figure 4) whereas in Africa rates may be 40È or higher (1, 1v0). Some

other explanations for the low prevalence rates include; recent introduction of the virus to the

population, recorded infections confined to high risk groups and early aggressive response to

the disease (1). An exception to the high rates in Africa is Chad where there is a higher

consumption of algae and prevalence rates of about 2.vÈ-v.6È have been reported. Other

locations in Africa showing lower prevalence rates include Niger and Senegal though there is no

data for whether there is a higher consumption of algae in these areas (141). Extracts from

seaweeds and Spirulina a cyanobacteria have been shown in vitro to inhibit a variety of

v4
enveloped viruses including HIV (1v6, 151). Its action seems to be mediated by direct

interaction with the HIV binding site of the CD4 receptor (v).

Algae are one of the first sources of natural compounds with in vitro anti-HIV activity.

They are grouped into six main classes based mainly on color. These classes are Chlorophyceae

(green algae), Phaeophyceae (brown algae), Pyrrophyceae (dinoflagellates), Chrysophyceae

(golden-brown algae), Bacillariophyceae (diatoms) and Rhodophyceae (red algae) (60).

Antibacterial activity of these plants has been reported as early as 1940 (125). Compounds

isolated from algae include steroids and sulfoglycolipids, but most research has centered on

natural and synthetic sulfated polysaccharides. These polysaccharides are able to block HIV

replication in cell culture in a manner that is dependent upon molecular weight and degree of

sulfation (125). They are found on the external surface of cell membranes, in pericellular

locations, in the extracellular matrix and at the mucous membrane. Their primary functions

seem to be cell to cell communication, to act as a barrier between tissues, cell adhesion, as a

reservoir for growth factors and also to protect against pathogens (1v8). Though they have

been studied primarily in blue green algae, extracts from over 60 species including brown and

red algae have been tested (145). Fucoidans, another group of sulfated polysaccharides, found

in brown seaweed stimulate immunoreactions, both humoral and cellular, and enhance

macrophage phagocytosis (118).

v5
Polysaccharide ID 
CC50 (ʅ ) C50 (ʅ ) 
C 0 (ʅ ) SI
GLP  10 
0.002 -.0001 
0.v6 -.025 5000

G P 10 
0.006 -.0006 0.vv-.01   
166
GLP A   
.6±0.1 0.01-.0004 
0.8 -.045 60

G PA 10 0.006-.0005 0.54-.0v 
166
DS 
. ±0.18 0.001-.0006 
0.1 -.00   00

% "0'    %31.%32)      %35
 ,%31    %32)       %350' $10'
 2 ,631    632      635 $   
  .%34%3  )     0' 
(145)

) &?.  +   

Carbohydrate-binding proteins (CBP), ~ 8-50 kDa proteins, can also be isolated from

algae, specifically from blue green algae, as well as sea corals, plants, invertebrates and

vertebrates (8). These show anti-HIV activity in cell culture perhaps due in some way to their

interacting with glycoproteins found on the viral envelop (8). Sulfated polysaccharides function

in the algae to maintain homeostasis in alkaline environments. It is thought that ingested

sulfated polysaccharides exert their influence by providing direct inhibition of viral uptake in

the gut, and also by immune enhancement such as increased interleukin-12 and interferon-1a

production, and stimulation of NK and B cells (1v6). Gut associated lymphoid tissue (GALT)

takes up large molecules which are presented to local immune cells and then may be

v6
transported to the lymph (1v6). The gut contains the majority of the body͛s lymphocytes and

as it an important route for HIV entry and potentially dominant site of HIV replication it is

important that we examine this (98).

Antimicrobicides aim to slow the spread of the disease by reducing transmission. As

most transmission of HIV occurs at the mucosal surfaces of the cervicovaginal and

gastrointestinal tracts, topical antimicrobicides can be applied to prevent entry (144). One

molecule that has shown promising results in vitro at preventing HIV entry is

cyclotriazadisulfonamide (CADA). It functions by decreasing the amount of the cell surface CD4

receptor which is necessary for viral entry (144). While this is not necessarily a treatment it

could be useful to examine the mechanisms behind this in order to provide more reliable

therapies.

Some people are homozygous for a v2 nucleotide deletion in the gene coding for the co-

receptor CCR5. This is naturally occurring in about 1È of the Caucasian population (6). The

CCR5 ɷv2 mutation confers some degree of immunity to HIV without significant immunological

disruption. A stem cell transplant from a person with this mutation has been shown to return

CD4+ T cell counts to normal in a patient with acute myeloid leukemia (65). These transplants

carry significant risks as they require the elimination of the patients͛ immune system (82).

Methods that could confer this mutation without having to rebuild the immune system would

carry much less risk. Research is being done into a CCR5 receptor antagonist. One such therapy

is to utilize modified forms of CCL5/RANTES (see below) that competitively bind gp120,

v7
reducing binding to CCR5 (6). It is important to note that the T-tropic (X4) strain of the virus, as

it uses CXCR4 as a co-receptor, is not affected by this deletion.

Despite treatment, progression of the disease eventually leads to AIDS. This is defined

by demonstrating a CD4+ T cell count of less than 200 cells per mmv or being diagnosed with

certain characteristic infections. AIDS defining infections include; AIDS dementia complex,

candida, coccifiocomycosis, cryptosporidiosis, cytomegalovirus, herpes simplex, histoplasmosis,

invasive cervical carcinoma, isospora beli, Kaposi͛s sarcoma, lymphoma, mycobacterial

infection, Pneumocystis jiroveci, pneumonia, progressive multifocal leukoencephalopathy,

salmonella, sepsis and wasting due to infection (92). Decay of the immune system is

continuous from here and patients eventually die.

- 



 Multiple studies have been done on people who appear to have some resistance to HIV

infection. These people typically fall into one of two groups; serologically discordant couples

and sex workers. Serologically discordant couple is a term that means that one partner is

positive for infection and the other is not (76). There are many terms used to described these

individual some of these include exposed seronegative (ESN), highly exposed persistently

seronegative (HEPS), and exposed uninfected EU (76). By studying these individuals insight into

not only the infectivity of the virus but also clues as to how to combat it can be gained. Some

of the factors that have been shown to be involved in this immunity include chemokine

receptor defects and various soluble suppressor factors (76).

v8
It was initially believed that the CD4 receptor was all that was required for virus entry

into the cell (92). After observing the slow progression of the disease in a certain population of

patients, it was discovered that a deletion in the gene responsible for chemokine receptors

CCR5 (ɷv2 mutation) rendered them less susceptible to the disease (92). This mutation results

in a nonfunctional chemokine receptor. As binding to both the CD4 receptor and a chemokine

receptor is required for viral entry this blocks the entry of the M-tropic (R5) strain of the virus.

As mentioned earlier this mutation is present in ~1È of the Caucasian population (6).

It has also been shown that in some groups despite repeated exposure to the virus there

was no infection (21, 59, 62, 6v, 66, 70). As sexual transmission of the virus is inefficient,

(99.5È of exposures do not result in infection), any method that decreases this could be highly

beneficial (66). This is important in that studying how these groups resist infection could lead

to a vaccination and/or antimicrobicides that will work to prevent infection. After ruling out

the CCR5 ɷv2 mutation as a cause of this resistance, it was observed that the mucosal layer

played a role in this resistance (21). It appears to have some protective factors such as HIV

specific IgA, cellular immune responses, and inhibitory factors like the protein RANTES

(regulated on activation, normal T expressed and secreted), SLP1 (synaptotagmin-like protein1),

alpha/beta-defensins, lysozyme, lactoferrin, calprotectin, cystatin, and histone H2A (21, 81).

Higher levels of IFN-ɲ secretion which interferes with viral infection of cells was also noted in

some HEPS (104).

While these mucosal factors have some inhibitory effects on HIV, that is not their

primary function. RANTES is a 68 amino acid chemokine that recruits leukocytes to the site of

v9
inflammation, activates lymphocytes and neutrophils. In HIV it can suppress viral replication in

vitro (5). SLP1 is in the family of carboxy terminal (C) -type tandem C2 proteins; they are

thought to be involved with vesicle transport and regulated endocytosis (47). Defensins were

discussed above. Lysozyme is an antimicrobial polypeptide found in the granules of neutrophils

and attacks the cell walls of bacteria (1v9). Lactoferrin interacts with reactive oxygen

intermediates (ROI) to facilitate the generation of hydroxyl radicals (1v9). It is a member of the

transferrin family of non-heme iron transporters and functions as an opsonin to increase

immune response (147). Cystatins are known to inhibit cysteine proteinases and may be

involved in targeting antimicrobial polypeptides (152).

) 7Ë0'   


 )  33&

 
 80 h) / 180h2

Another study found that a small molecular weight protein elafin/trappin-2 was

elevated in secretions from resistant women. This protein is also called skin-derived

40
antileukoproteinase (SKALP) (66). This protein is also related to the SLP1 found in the previous

study to be elevated in resistant individuals.

)


In 2005 there were 4.5 million new infections and 2.8 million deaths worldwide (128).

Infection rates have skewed to include more women and increasingly, new infections are

diagnosed in those under 25 (between 25-40È) (1). Sub-Saharan Africa continues to have a

disproportionately large number of infected (table 6) and 70È of those are women. This has to

do with not only sexual mores but also the fact that male to female transmission rates are

higher than female to male rates. Transmission risk varies with viral load, infection stage,

treatment status, circumcision status, presence of other sexually transmitted infections, and

social behavior (1).

Geographic Area Number infected


North America 1.v million
Caribbean vv0,000
Latin America 1.6 million
Western and 720,000
Central Europe
N. Africa & Middle 440,000
east
Sub-Saharan Africa 24.5 million
Eastern Europe & 1.5 million
Central Asia
East Asia 680,000
South and South 7.6 million
East Asia
Oceania 780,000
&Ë0'  33%  
 
(1)

41
   4   

There are many more mechanism of immunity that are potentially interesting to explore

further. Mechanisms to inhibit, deactivate, or block the CCR5 receptors would make for an

interesting course of study as a mechanism to reduce infection. Also of interest would be a

mechanism that would allow the preferential blocking of the CXCR4 co-receptor only where it

interacts with HIV would make for a viable combined therapy. As the structure of these cell

surface glycoproteins gp120 and gp41 is studied perhaps drugs can be developed to interfere

with the binding of these structures thus blocking viral entry. More study should be given to

the role of ɽ-defensins in the long term control of SIV. SIV rarely if ever progresses into full

blown AIDS and this may be a function of defective CCR5 receptors but it could also be related

to the role of retrocyclin.

Further investigation into why consuming sulfated polysaccharides is beneficial would

also be a good avenue of study. In places where access to care is unavailable or limited, being

able to control infection by alternate/non-drug means would be invaluable. Sulfated

polysaccharides are also important as they are the only mechanism that has been shown to

prevent HIV entry into cells of the gut. As this is an important area of viral replication these

could play a key role in limiting the infectivity of the virus.

42
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,**,:*9
,*Ë , *.*2007. Global Epidemiology of
HIV-AIDS. Infect. Dis. Clin. North Am. ;1-17.

2. ,*,*<  ,<*!,-*(,9*(,  *8*2002. Molecular biology


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v. 
  ,*,*,*
 ,*8,0* ,(*! ,*<*8 , =**9
*
1999. A new sulfated ɴ-galactan from clams with anti-HIV activity. Carbohydr. Res. #;121-
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4.  ,*,*,*9
 , ?**  *2005. Distribution of
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