[Emerging Infectious Diseases * Volume 3 * Number 3 * July - September 1997]

Perspectives

Host Genes and HIV: The Role of the Chemokine Receptor Gene CCR5 and Its
Allele ([Image of delta] 32 CCR5)

Janet M. McNicholl, Dawn K. Smith, Shoukat H. Qari, and Thomas Hodge
Centers for Disease Control and Prevention, Atlanta, Georgia, USA
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      Since the late 1970s, 8.4 million people worldwide, including
      1.7 million children, have died of AIDS, and an estimated 22
      million people are infected with human immunodeficiency virus
      (HIV)(1). During 1995 and 1996, major clinical and laboratory
      discoveries regarding HIV pathogenesis provided new hope for
      the prevention and treatment of HIV infection. One major
      discovery was that members of the chemokine receptor family
      serve as cofactors for HIV entry into cells. We describe the
      role of allelic polymorphism in the gene coding for the CCR5
      chemokine receptor with regard to susceptibility to and disease
      course of HIV infection. We also examine the effect of this
      discovery on medical and public health practices.

The HIV Epidemic: A Global Problem

The World Health Organization has estimated that 22.6 million people are
infected with human immunodeficiency virus (HIV) as of mid-1996 (1). The
highest prevalence of HIV occurs in parts of Asia and sub-Saharan Africa.
In 1995, HIV-associated illnesses caused the deaths of 1.3 million people,
including 300,000 children under 5 years of age. No protective vaccine or
cure is available, and while prevention methods are reducing incidence in
some countries, HIV disease is expected to increase. Earlier HIV infection
diagnosis, inhibition of ongoing HIV replication with antiretroviral
therapy (in industrialized countries), and prevention and treatment of
opportunistic infections and cancers delay the onset of AIDS and increase
the life expectancy of HIV-infected persons.

An Overview of Host Genes and HIV Infection

Since the early years of the HIV epidemic, significant differences in the
rate of disease progression have been observed in longitudinally followed
HIV-infected persons. The role of genes of the human leukocyte antigen
(HLA) system in determining the course of disease has been examined by
using such measures as the CD4+ cell count or the length of time between
HIV infection and AIDS (2-6). Several HLA genes or haplotypes appear to
influence disease progression, although the effects are complex and may
depend on interactions with other host genes.

Studies of the effect of host genes on susceptibility to HIV infection were
facilitated by the identification of persons who were persistently exposed
to HIV but remained uninfected (7-12). Before the discovery of the role of
chemokine receptor gene polymorphism in HIV infection, only genes of the
HLA system were thought to protect against HIV infection. For example,
certain distributions of HLA class I alleles were observed in uninfected
female commercial sex workers in Africa (13,14) and Thailand (11), who had
been highly exposed to HIV; additional class I and II alleles that may be
associated with remaining uninfected have been identified (6). Several
mechanisms, some related to cytotoxic T-cell function, have been suggested
to explain these findings (7,10,11,15).

Non-HLA genetic factors also influence susceptibility to HIV infection and
the course of HIV disease. In 1996 and 1997, studies confirmed the
protective role of homozygosity for a 32 base pair (bp) deletion in the
chemokine receptor gene, CCR5 ([Image of delta]32 CCR5), against HIV infection
(9,12,16-19). Until recent reports of HIV infection in three persons
homozygous for the 32 bp CCR5 deletion (20-22), no cases of HIV infection
had been reported in studies of more than 60 persons homozygous for the
CCR5 32 bp deletion. Presence of one copy of the deleted CCR5 gene also
influences the course of disease as the onset of AIDS occurs later for some
heterozygous persons than for those homozygous for the wild type CCR5
(12,17-19,23). The discovery of the role of CCR5 alleles has prompted
studies of the possible role of many other host genes in HIV infection
(24-26).

What Are Chemokine Receptors?

Chemokine receptors are cell surface proteins that bind small peptides
called chemokines (27). Chemokines can be classified into three groups
based on the number and location of conserved cysteines: C, CC, and CXC.
Chemokine receptors are grouped into families on the basis of the chemokine
ligands they bind: CC, CXC, or both (27; Table 1). Some receptors are
promiscuous, while others are selective in terms of ligand binding. The
receptors are widely distributed on hematopoietic and other cells, but the
Duffy antigen of erythrocytes (DARC) is the only member expressed on cells
of erythroid lineage. Several human chemokine receptors have been
classified as such on the basis of similarity of gene sequences and
predicted protein structures, but their ligands have not been identified
(orphan receptors). Among these is the recently identified TER1 gene (28).

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Table 1. Human chemokine receptors.

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Receptors                         Predominant                Chromo-
(Old                              expression/      Pathogens some     GenBank
names)                            Tissue           (super-   location Acc.#
(super                            distribution     script b)
script a)         Ligands
           ---------------------
              C-C       CXC
           chemokines chemokines
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CC
Receptors
---------
CCR1       MIP-1(alpha)           monocytes, T                3p21    L10918
 (CC CKR1) (beta), MCP-3          cells

CCR2A      MCP-1                  T cells, basophils          3p21    U03882
 (MCP-1Ra)                        monocytes

CCR2B      MCP-1,3,4              HIV-1                       (NSI)   U03905
 (MCP-1Rb)

CCR3       EotTaxin               eosinphil, baso-   HIV-1    3P21    U28694
 (CKR3)    RANES,                 phils, microglial  (NSI)
           MCP-2,3,4              cells, and
                                  possibly monocytes;
                                  little expression
                                  in peripheral
                                  blood T-lymphocytes
                                  or dendritic cells

CCR4       TARC                   basophils, T cells          3P24    X85740

CCR5       RANTES                 monocytes,         HIV-1    3P21    U57840
(superscript c)                   dendritic cells,   (NSI)HIV-2
 (CC CKR5) MIP-1(symbol           microglial cells
           alpha),                T cells
           (symbol beta)

CXC Receptors
--------------
CXCR1                  IL-8       neutrophils, NK             2q35    M68932
 (IL-8 RA)                        cells

CXCR2                  IL-8, MGSA                                     M73969
 (IL-8 RB)             gro-(symbol
                       alpha),
                       NAP-2, IP-10
                       ENA-78, Mig

CXCR3                  IP-10, Mig activated T cells                   X95876

CXCR4                  SDF-1      wide: CD4+ and CD4- HIV-1   2q21    M99293
 (Fusin, LESTR,                   cells, monocytes,    (SI)
  HUMSTR)                         macrophages,        HIV-2
                                  dendritic cells,
                                  B cells, other
                                  tissues, e.g.,
                                  brain, lung, spleen

CC/CXC Receptor
---------------
DARC       RANTES,     IL-8,      endothelial cells,  Plasmo-  1      U01839
 (Duffy    MCP-1,      gro-       erythrocytes        dium
  antigen) TARC etc.   (symbol                        vivax
                       alpha) etc.

Others (superscript d)
----------------------
STRL33     ND          ND         lymphoid tissues    HIV-1    3      U73529
                                  and activated T cells

HCMV-US28  MIP-1                  fibroblasts         HIV-1   N/A     X17403
           (symbol                infected with CMV
           alpha),
           (symbol beta),
           RANTES

ChemR1     ND          ND         T lymphocytes,              3p21-24 Y08456
                                  polymorphonuclear
                                  cells

CMKBRL1    ND          ND         neutrophils,                3p21    U28934
                                  monocytes, brain,
                                  liver, lung, skeletal
                                  muscles

TER1       ND          ND         thymus, spleen              3p21    U62556

V28        ND          ND         neural and lymphoid         3p21    U20350
                                  tissue

D2S201E    ND          ND         wide,including cells        2q21    M99293
                                  of hemopoietic origin

BLR1       ND          ND         B lymphocytes                       X68149

EBI1       ND          ND         B lymphocytes                       L08176

GBR1,2,5   ND          ND                                ND           L36149
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(superscript a)New nomenclature for CC and CXC chemokine receptors was
adopted at the Gordon Research Conference on Chemotactic Cytokines, June
23-28, 1996. (superscript b)Pathogens using this receptor for infection.
(superscript c)>The 32bp deleted allele of CCR5 has been referred to as
CCR5-2 (19). (superscript d) Chemokine receptor-like genes whose predicted
proteins have 7 transmembrane domains.

Abbreviations: BLR1, Burkitt=s lymphoma receptor-1; CMKBRL1, Chemokine b
receptor like-1; DARC, duffy antigen/receptor for chemokines; EBI1,
Epstein-Barr virus-induced receptor; ENA78, epithelial-derived
neutrophil-activating peptide-78; GPR, G protein coupled receptor; gro,
growth related gene product; HCMV, human cytomegalovirus; HUMSTR, human
serum transmembrane segment receptor; IL, interleukin; IP-10,
interferon-gamma inducible 10kD protein; LESTR, leukocyte-expressed
seven-transmembrane-domain receptor; MCP, monocyte chemotactic protein;
Mig, monokine induced by interferon gamma; MIP, macrophage inflammatory
protein; NSI, non-syncytium inducing; N/A, not applicable; NAP-2,
neutrophil-activating protein-2; ND, not determined; RANTES, regulated on
activation, normal T cell expressed and secreted; SDF-1, stromal
cell-derived factor-1; STRL33, seven transmembrane-domain receptor from
lymphocyte clone 33;TARC, thymus and activation regulated chemokine.

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The characteristic feature of all chemokine receptors is a serpentine 7
transmembrane-spanning domain structure, which is shared with other
receptors; e.g., the rhodopsin and the thyrotrophin receptors (Figure 1).
Extracellular portions are involved in chemokine binding, while
intracellular portions are involved in cell signaling. The effect of
receptor-ligand interactions is usually mediated through G-protein coupled
interactions; results in alterations in cell function such as activation,
motion, or migration, usually along a chemokine concentration gradient; and
varies depending on the chemokine bound and the cell type (27).

Some chemokine receptors have a role in infectious disease susceptibility
or pathogenesis. Several of the CC or CXC receptors are used by HIV-1 or
HIV-2 as entry cofactors (Table 1). DARC serves as a cofactor for entry of
Plasmodium vivax into erythrocytes (29); and as is the case with the
32bp-deleted CCR5 and HIV, resistance to P. vivax malaria is associated
with lack of DARC expression (29), probably due to polymorphism of the DARC
gene promoter (30). Several viruses (Epstein-Barr virus, cytomegalovirus,
and Herpes virus samiri) contain functional homologues of human chemokine
receptors, which suggests that the viruses may use these receptors to
subvert the effects of host chemokines (27). They may also serve as HIV
entry cofactors for human cells, as observed for HCMV-US28 (31).

Chemokine Receptors and HIV Infection

The first clue that chemokine-related events were important in HIV
pathogenesis came from work in R. Gallo's laboratory, which showed that
high levels of chemokines could inhibit HIV replication in vitro (32). Then
a cohort of highly exposed, HIV-negative men had high circulating levels of
several chemokines, such as RANTES, MIP-1[Image of alpha] , and MIP-1 (8). These
data led to the hypothesis that chemokines might prevent HIV infection by
binding to the elusive HIV entry cofactor. Since the discovery of CD4 in
the 1980s as an HIV receptor, it had become apparent that other factors
were required for HIV to enter cells. For example, mouse cells expressing
CD4 could not be infected with HIV (33). Using a novel cloning strategy, E.
Berger and colleagues first demonstrated in May 1996 that CXCR4, a
chemokine receptor for which no ligand had yet been determined, was an
entry cofactor for T-cell tropic or syncytium-inducing (SI) viruses (34).
Shortly before this, Samson et al. had cloned CCR5 (then known as CC-CKR5)
and shown that its ligands included RANTES, MIP-1a, and MIP-1 (35). These
data encouraged several groups to examine the role of CCR5 in HIV entry,
and within weeks of the CXCR4 publication, five additional publications
reported CCR5 and CCR3 as HIV entry cofactors for macrophage-tropic or
non-syncytium-inducing (NSI) viruses (33,36-39) and CCR3 and CCR2b as entry
factors for dual tropic HIVs (38,39). Recent studies of brain-derived
microglial cells have shown that CCR5 and CCR3 permit entry of HIV into
these cells (40).
[Image]
Figures Not Available in Ascii

Figure 1. Predicted structure and amino acid sequence of CCR5. The typical
serpentine structure is depicted with three extracellular (top) and three
intracellular (bottom) loops and seven transmembrane (TM) domains. The
shaded horizontal band represents the cell membrane. Amino acids are listed
with a single letter code. Residues that are identical to those of CCR2b
are indicated by dark shading, and highly conservative substitutions are
indicated by light shading. Extracellular cysteine residues are indicated
by bars, and the single N-linked glycosylation consensus site is indicated
by an asterisk. Reprinted and modified with permission from the authors and
Cell (39). Copyright (1996) Cell Press.

The tropism of HIV strains appears to be determined in part by the way they
use chemokine receptors (Table 1, Figure 2). In peripheral blood, although
both T cells and monocytes express CXCR4 and CCR5 (41), HIV strains that
use CXCR4 tend to infect predominantly T cells, while strains that use CCR5
infect T cells and monocytes. This tropism correlates with the ability of
the virus to induce syncytia in T-cell lines. SI viruses (often present
late in the course of HIV infection), tend to be CXCR4 tropic, while NSI
viruses (which appear to be the viruses transmitted in vivo) tend to be
CCR5 tropic (33,34,36-39). However, some primary SI viruses can use either
CXCR4 or CCR5 (42), and some may use both coreceptors, as well as CCR2b or
CCR3 (39). When classified based on chemokine receptor use, HIV types are
independent of their genetic relatedness (43).
[Image]
Figures Not Available in Ascii
Figure 2. Chemokine receptors and cell tropism of HIV. Three cell types are
illustrated, an in vitro passaged T-cell line (Tl), a monocyte/macrophage
(M), and a circulating T-cell (T). T-cell lines express CXCR4 but not CCR5;
macrophages and circulating peripheral blood T-cells express both
receptors, although the amounts of CXCR4 are lower on macrophages (as
indicated by the small CXCR4 symbol). M-tropic HIV, because of certain
envelope amino acid sequ
ences, binds to CCR5 and can enter both macrophages
and circulating T cells. T-tropic HIV preferentially binds CXCR4 and enters
T cells or T-cell lines. After binding to the chemokine receptor and to
CD4, the viruses enter by fusion with the cell membrane. Cells with the
32bp deleted form of CCR5 do not express cell surface CCR5, and, although
M-tropic HIV can bind CD4, it cannot enter the cell. If the cells express
CXCR4, they can still be infected with T-tropic viruses (not shown). Note
that this figure does not depict the actual size relationships of the
proteins, cells, or viruses.

The exact mechanism by which HIV interacts with CXCR4 or CCR5 and by which
this interaction, together with CD4 binding, leads to virus entry has not
been clearly defined (44). It appears to involve the interactions of the V3
loop and other parts of the outer envelope protein gp120 (45-47) with
extracellular domains of CCR5 or CCR2b (48,49) and may involve multistep
interactions with CD4, the chemokine receptor, and other cell surface
components (50). Chemokines may inhibit HIV entry through receptor
blockade, desensitization, sequestration, or internalization; through
alterations in receptor affinity; or by inhibiting postbinding steps (51),
such as phosphorylation, through G-coupled mechanisms.

Chemokine Receptor Gene Polymorphism and Susceptibility to HIV Infection

In 1995 and 1996, researchers at the Aaron Diamond AIDS Research Center
began studies of highly HIV-exposed seronegative men whose cells produced
high levels of chemokines in vitro (8,52,). Cells from several of these men
were not infectable with primary or NSI type viruses, but were infectable
with SI viruses (52). These data suggested some protective mechanism
related to the CCR5 rather than to the CXCR4 receptor because of the
chemokine ligand profile of the CCR5 receptor (Table 1) and stimulated
studies that led to the cloning and restriction digestion of CCR5 in these
persons (52). The novel deletion of 32bp in CCR5 reported by Liu et al. (9)
in August 1996, to be present in three of the 15 HIV-exposed but
seronegative men was also independently reported in September by Samson et
al. (16) and Dean et al. (17; Table 2). In these studies, the distribution
of homozygosity and heterozygosity of the deleted allele, in small and
large cross-sectional or prospective studies of HIV-unexposed, -exposed, or
-infected persons (mainly European or North American Caucasians) strongly
supported the protective effect of the deletion. Since these initial
observations, several studies of other populations have reported the same
finding: an enrichment of the homozygous [Image of delta]32/[Image of delta]32 CCR5 genotype
in highly HIV-exposed persons who remain uninfected (12,18,19; Table 2). In
persons with well-documented HIV exposure, the prevalence of homozygosity
for the [Image of delta]32/[Image of delta]32 CCR5 genotype increases with increasing HIV
exposure (12), the wild type and [Image of delta]32 CCR5 alleles (referred to by
some as CCR5-1 and CCR5-2 [19]) can be easily detected by polymerase chain
reaction (PCR) techniques, with or without enzymatic digestion
(9,12,16,17), or more recently, by heteroduplex studies (19). An example of
the results of a PCR followed by restriction digestion is given in Figure
3. Persons who are homozygous for the wild type CCR5 (W) or the allele with
the 32 bp deletion ([Image of delta]32) or heterozygous for both alleles can be
easily distinguished because the deleted fragment reduces the size of the
amplified product (Figure 3).
[Image]
Figures Not Available in Ascii
Figure 3. Differentiation of CCR5 genotypes by gel electrophoresis. Band
patterns of persons with homozygous wild type (W/W), homozygous 32 bp
deletion ([Image of delta]32/[Image of delta]32) or heterozygous W/[Image of delta]32 CCR5 genotypes
are shown. PCR amplification of the C-terminal of the CCR5 gene, subsequent
digestion with the EcoRI restriction enzyme, and agarose gel
electrophoresis of the digested DNA yield a 182 bp band for the wild type
CCR5 gene, a 150 bp band for the 32 allele, and both bands in the case of a
heterozygous person.

The 32 bp deletion occurs at a site of a repeat motif in the CCR5 gene (16;
Figure 4) and results in a frame shift in the coding sequence that produces
a defective CCR5, which is not expressed on the surface of cells (Figures 2
and 5). The deletion results in the loss of three of the seven
transmembrane domains, two of the three outer loops, and the intracellular
signaling domain (Figure 5).

Does the CCR5 32 Bp Deletion Provide Absolute Protection Against HIV
Infection?

The genetic link to remaining HIV-negative in spite of continued HIV
exposure was the subject of much discussion during the summer and fall of
1996. Was the protection absolute? Who should be tested for the gene? Would
persons told they were homozygous for the deletion engage in high-risk
behavior more frequently? Although at that point no one homozygous for the
deletion had been found HIV-positive, the fact that CXCR4 gene and several
other chemokine receptors could mediate HIV entry suggested caution in
assuming that persons with the CCR5 [Image of delta]32/[Image of delta]32 genotype would be
absolutely resistant to HIV infection. Since then, three HIV-infected
persons with this genotype have been reported (20-22; Table 2); for one,
the mode of transmission was most likely blood products (21), while the
only reported exposure risk for the other two was homosexual behavior
(20,22). Sequences isolated from these persons suggested the SI virus
phenotype, i.e., viruses that most likely used CXCR4 as entry cofactors
(21; R. Ffrench, pers. comm.). As several other non-CCR5 receptors (CCR3
and CCR2b) are also HIV entry cofactors, even non-SI viruses might be
transmissible in vivo in persons homozygous for the CCR5 32 bp deletion.
Thus, HIV entry by non-CCR5 dependent mechanisms can occur in vivo, and
persons who have the CCR5 [Image of delta]32/[Image of delta]32 genotype are not absolutely
protected.
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Table 2. Population studies of CCR5 genotypes
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 Author           Population        Ethnicity        HIV     No.  (superscript a)CCR5 Genotypes           [Image of
                                                     Status       % (N)                                   delta] 32
                                                                                                          allele
                                                                                                          frequency
                                                                                                          (%)

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                                                                  W/W           w/[Image  [Image of
                                                                                of        delta]32/[Image
                                                                                delta]32  of delta]

 Liu              High-risk, USA    Caucasian        Neg     15   80.0          0 (0)     20.0 (3)        0.200
 et al. (9)                                                       (12)

                  General, USA      Caucasian        Neg     122  80.3          19.7      0 (0)           0.098
                                                                  (98)          (24)
                  General, S.       Venezuelan       Neg     46   100           0 (0)     0 (0)           0
                  America                                         (46)
 Samson           General, Europe   Caucasian        Neg     704  82.7          16.2      1.1 (8)         0.092
 et al. (16)                                                      (582)         (114)
                  General, Asia     Japanese         Neg     248  100           0 (0)     0 (0)           0
                                                                  (248)
                  General, Africa   Ctrl/West        Neg     124  100           0 (0)     0 (0)           0
                                    African                       (124)
                  AIDS clinics,     Caucasian        Pos     723  89.2          10.8      0 (0)           0.054
                  Paris                                           (645)         (78)
 Dean             High-risk, USA    Mixed            Neg     883  81.9          15.6      2.4             0.10
 et                                                               (724)         (138)     (21)
 al.(superscript
 b)(17)
                  High-risk, USA    Mixed            Pos     1883 85.9          14.0      0.0005          0.071
                                                                  (1618)        (264)     (1)
                  High-risk, USA    African-Amer     Mixed   620  96.6          3.4       0 (0)           0.017
                                                                  (599)         (21)
                  High-risk, USA    Caucasian        Mixed   1250                                         0.115
                  Low-risk, USA     Caucasian        Mixed   143                                          0.080
 Huang            High-risk, USA    Caucasian        Pos     461  79.8          20.2      0 (0)           0.101
 et al.(12)                                                       (368)         (93)
                  High-risk, USA    Caucasian        Neg     446  78.0          16.7      3.6             0.128
                                                                  (348)         (82)
                  Blood donors, USA Caucasian        Neg     637  85.2          13.3      1.4             0.08
                                    (95%)
                  General,          Black            Mixed   137  100           0 (0)     0 (0)           0
                  Africa/Haiti
                  General, Asia     China/Thai/other Mixed   191  100           0 (0)     0 (0)           0
 Michael          High-risk, USA    Mixed            Pos     406  87.5          14.3      0 (0)           0.071
 et al. (18)                                                      (348)         (58)
                  High-risk, USA    Mixed            Neg     21   71.4          9.5       19.1            0.238
                                                                  (15)          (2)       (4)
                  Intermediate-risk,Mixed            Neg     240  78.3          20.4      1.3             0.115
                  USA                                             (188)         (49)      (3)
 Biti             High-risk,        Caucasian        Pos     265  N/A           N/A       0.004           N/A
 et al.(20)       Australia                                       (superscript            (1)
                                                                  c)
 Theodoru         High-risk, Europe                  Pos     412  N/A           N/A       0.002           N/A
 et al. (22)                                                                              (1)
 Eugenolsen       High-risk, DenmarkCaucasian        Neg     35   74            20        6               0.157
 et al. (23)                                                      (26)          (7)       (2)
                  High-risk, DenmarkCaucasian        Pos     99   78            22        0               0.111
                                                                  (77)          (22)
                  Blood donors,     Caucasian        Neg     37   73            24        3               0.149
                  Denmark                                         (27)          (9)       (1)
 Zimmerman        Blood donors,     Caucasian        Neg     387  77.5          21.7      0.8             0.116
 et al. (19)      North America                                   (300)         (84)      (3)
                                    African Amer     Neg     294  94.2          5.8       0 (0)           0.29
                                                                  (277)         (17)
                                    Hispanic         Neg     290  92.8          6.9       0.3             0.38
                                                                  (269)         (20)      (1)
                                    Asian            Neg     164  99.4          0.6       0 (0)           0.003
                                                                  (163)         (1)
                                    Native Amer.     Neg     87   83.9          12.6      3.4             0.098
                                                                  (73)          (11)      (3)
                  Blood donors,     Tamil            Neg     46   100           0 (0)     0(0)            0
                  India                                           (46)
                  Blood donors, W.  Black            Neg     40   100           0 (0)     0 (0)           0
                  Africa                                          (40)
                  High Risk, USA    Caucasian        Pos     614  77.4          22.6      0 (0)           0.113
                                                                  (475)         (139)
                  High Risk, USA    African Amer     Pos     86   97.7          2.3       0 (0)           0.012
                                                                  (84)          (2)
                  High Risk, USA    Hispanic         Pos     45   93.3          6.7       0 (0)           0.033
                                                                  (42)          (3)
                  High Risk USA     Caucasian        Neg     111  73.9          21.6      4.5             0.0153
                                                                  (82)          (24)      (5)
                  High Risk, USA    African Amer     Neg     2    50.0          0 (0)     0.250
                                                                  (1)
                  High Risk, USA    Hispanic         Neg     12   91.7          8.3       0 (0)           0.042
                                                                  (11)          (1)

(superscript a)CCR5 genotypes: W/W, homozygous wild type; W/[Image of
delta] 32, heterozygous wild type/32bp deletion; [Image of delta]32/[Image
of delta]32, homozygous for the 32 bp deletion.

(superscript b) Numbers updated to include additional Multicenter
Hemophiulia Cohort Study patients studied by O'Brien et al. (21) and Dean
et al. (pers .comm.).

(superscript c) Not available
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[Image]
Figures Not Available in Ascii
Figure 4. Partial CCR5 gene and amino acid sequence with 32 bp deletion.
Nucleotide sequence of the CCR5 gene surrounding the deleted region, and
translation into the normal receptor (top lines) or the truncated mutant (
[Image of delta] 32 CCR5, bottom lines). The 10-bp direct repeat is represented in
bold italics and the deleted nucleotides are represented in noncapitalized
font.

[Image]
Figures Not Available in Ascii
Figure 5. Predicted structure and amino acid sequence of the mutant form of
human CCR5. The mutant protein lacks the last three transmembrane segments
of CCR5, as well as the regions involved in G-protein coupling. The
transmembrane organization is given by analogy with the predicted
transmembrane structure of the wild-type CCR5, although the correct
maturation of the mutant protein up to the plasma membrane has not been
demonstrated. The shaded horizontal band represents membrane (s) of
intracellular organelles. Amino acids represented in italic and dark
shading correspond to unnatural residues resulting from the frame shift
caused by the deletion. Figures 4 and 5 reprinted and modified with
permission from the authors and Nature (16) Copyright (1996) Macmillan
Magazines Ltd.

Doe
s CCR5 Gene Polymorphism Determine All Resistance to HIV?

The highest reported prevalence of homozygosity for the [Image of delta]32 CCR5
allele deletion in persons highly exposed to HIV but uninfected is 33%
(9,12). However, most (96% according to one estimate [19]) highly exposed
HIV-seronegative persons are not homozygous for the [Image of delta]32 CCR5 allele.
Moreover, among exposed but persistently HIV-seronegative persons in Africa
and Thailand, no one positive for the [Image of delta]32 CCR5 allele has been found
(CDC unpub. data; S. Rowland-Jones, F. Plummer, pers. comm.). These data
suggest that many mechanisms may contribute to remaining seronegative
despite high HIV exposure: chemokine-related mechanisms, such as altered
CCR5 levels on the cell surface (53) or the level of circulating
chemokines; other immune system genes, such as those of the HLA system
(10,11,14); and immune responses, such as those of cytotoxic T cells
(7,15). The relative importance of these mechanisms in populations with
differing modes of exposure or genetic backgrounds needs to be elucidated.

CCR5 Polymorphism and Its Interaction with Other Factors in the Course of
HIV Disease

Several large studies of the effect of CCR5 genotypes on the course of HIV
disease have been published (Table 2). Dean et al. examined the course of
HIV (time to AIDS) in several U. S. populations with different exposure to
HIV (homosexuals, intravenous drug users, persons with hemophilia) and
found that persons with one copy of the deleted CCR5 gene had a delayed
(approximately 2 years longer) progression to AIDS when compared with those
with the homozygous wild type genotype (17). Similar findings have been
reported (19,23) in Caucasian HIV-positive homosexual men. Another study of
HIV-positive homosexuals did not find such a striking effect of
heterozygosity, although progression to AIDS was again slowed (12). One
clue to the varying effects of the heterozygous CCR5 genotype on
progression to AIDS in different groups comes from another U. S. study of
male homosexuals, in which men with one copy of the [Image of delta]32 CCR5 genotype
had delayed progression to AIDS, particularly if their circulating viruses
were of the NSI phenotype (18). Paradoxically, once AIDS is diagnosed,
persons with one copy of [Image of delta]32 CCR5 may have a more rapid disease
course. Thus, the effect of the CCR5 genotype on disease progression may
depend on the phenotype and chemokine receptor use of circulating viruses.
The finding that microglial cells can be infected with HIV through CCR3- or
CCR5-dependent mechanisms (40) also suggests that the clinical spectrum of
HIV disease may also depend on the CCR5 genotype.1

Thus, complex interactions between virus and host chemokine receptor
genotype and virus and cell chemokine receptor phenotype exist. Studies to
elucidate the interaction of these genotypes with each other, with virus
phenotypes, and with other factors that influence the course of HIV disease
are ongoing (24,25). Preliminary data from the Multi Center AIDS Cohort
Study suggest that the effect of CCR5 and HLA genotype on survival or
disease course are independent and that the effect of HLA on disease
outcomes may be greater than the effect of CCR5 genotype (54).

Population Studies of CCR5 Genotypes

The distribution of the 32bp deletion in different populations is
summarized in Table 2. Among Caucasians in North America or Europe, the
prevalence of homozygosity for the 32 bp deletion is approximately 1%, and
10% to 20% are heterozygous. In smaller numbers of non-Caucasians,
heterozygosity is found in approximately 6% of African-Americans, 7% of
Hispanics, 13% of Native Americans, and fewer than 1% of Asians (17,19;
CDC, unpub. data). Several studies of non-Caucasian populations including
persons from parts of Africa (Zaire, Burkina Faso, Cameroon, Senegal,
Benin, Uganda, Rwanda, Kenya, Malawi, Tanzania, Sierre Leone) (12,16,19),
Haiti (12), parts of Asia (Thailand, India, China, Korea, Japan, the
Philippines) (12,16,19; CDC, unpub. data), and Venezuela (9) have not found
the 32 bp deletion among the HIV-infected or -uninfected persons tested
(Table 2). One additional study (of more than 3,000 persons) found similar
global distributions of CCR5 genotypes (and noted a decreasing frequency
from north to south in Europe and Asia).2

Other Chemokine Receptor Polymorphisms

Several other point mutations in CCR5 have been observed (17), but their
population prevalence and their role in HIV infection or disease
progression have not been reported. No polymorphisms in CXCR4 have been
reported to date. CCR3 has two alleles (S or T at position 276) (55-57),
and similarly, the population prevalence of these alleles and their role in
HIV infection have not yet been determined.

Implications for Management and Understanding of the Global HIV Epidemic

The discovery of the role of chemokine receptors as coreceptors, along with
CD4, for HIV entry has led to a burgeoning of public and private research
in HIV pathogenesis, new therapies, and new approaches to vaccines (52,58).
For example, in addition to the classification of HIV types on the basis of
genetic or neutralization phenotype relatedness, a new virus classification
made on the basis of their receptor use has emerged. New therapies based on
receptor mimics, receptor ligands, chemokines, or chemokine analogs are
being developed and tested in vitro. Genetic engineering with chemokine
receptors is being used to develop animal models for HIV transmission and
disease progression. And, in the arena of vaccines, a new concept has
developed: the induction of high levels of HIV-blocking chemokines such as
RANTES, MIP-1[Image of delta] , and MIP-1 as desirable properties of an HIV
vaccine.

New treatments and prevention measures for HIV raise ethical, social, and
legal issues that also relate to a number of other infectious and chronic
diseases (e.g., malaria, breast cancer, and diabetes) as new genes
influencing the risk for these diseases are discovered. Should all persons
at risk for HIV infection be tested for CCR5 gene polymorphism? Should all
persons homozygous for the [Image of delta]32 CCR5 genotype receive specific
counseling regarding their risk for HIV and other sexually transmitted
infections? If a person is heterozygous and infected with HIV, should HIV
treatment be altered? Could knowledge of one's genotype favorably or
unfavorably influence health insurance, life insurance, or employment
opportunities? As a result of these questions, educational materials have
become available for the public and public health professionals.3

Another finding that provoked discussion was the uneven distribution of the
genotype across geographic and racial groups. One hypothesis was that some
previous epidemic, restricted to Europe, had led to a survival advantage
among persons homozygous or heterozygous for the CCR5 [Image of delta]32 genotype
and had resulted in a concentration of the CCR5 [Image of delta]32 genotype in
persons of European ancestry. The "black death" of Northern Europe
(1347-1350) and other large epidemics have been used as examples (59,60).
Thus, certain populations appear to have an increased survival advantage in
the new HIV epidemic. Conversely, populations with lower prevalence of the
[Image of delta]32 CCR5 genotype might be expected to have a higher prevalence of
HIV infection or a more rapid course of the epidemic. In the United States,
a greater risk for HIV infection has been found among African-Americans
than among Caucasians, when known risk factors, including social class,
were controlled (61). The prevalence of the [Image of delta]32 CCR5 allele is lower
in African-Americans than in Caucasians. While increasing HIV prevalence in
parts of Asia and Africa may be attributed to social and demographic
factors, as well as differences in the phenotype of circulating viruses
(62), the racial distribution of HIV risk raises the possibility that
differences in the distribution of the [Image of delta]32 CCR5 allele or other
heritable host factors might influence the rate of transmission or the
speed of the epidemic in different racial groups.

Conclusions

These findings have defined a new role for a gene that normally controls
cell migration; have identified new alleles of the gene that determines
whether someone becomes infected with HIV and how the disease progresses
once infection has occurred; and have identified several other new
HIV-receptors that mediate viral entry into different cell types. The
findings have precipitated new classifications of HIV phenotypes by cell
tropism and receptor use and opened new areas for drug and vaccine
development. These findings have also enhanced other research: studies of
host factors affecting the acquisition and clinical course of infectious
diseases, the differential distribution of genetic characteristics in
populations and their potential effects on health status, the translation
of genetics research into public health measures, and the social
implications of genetic differences affecting illness and death caused by
infectious and other diseases.

About the Author

Dr. McNicholl is a scientist in the Immunology Branch, Division of AIDS,
STD, TB, and Laboratory Research, NCID, CDC, and an Assistant Professor of
Medicine (Rheumatology) and Pathology at Emory University. Her research
includes immunogenetics and T cell antigen recognition in infectious and
autoimmune diseases, and their application to vaccines. In collaboration
with other scientists at CDC and in Thailand, she is investigating
immunologic and genetic explanations for apparent resistance to HIV of
highly exposed persistently seronegative commercial sex workers in
Thailand. She also serves as the NCID coordinator for CDC genetics
activities.

Address for correspondence: Janet McNicholl, Centers for Disease Control
and Prevention, MS A-25, 1600 Clifton Rd. N.E., Atlanta, GA 30333 USA; fax:
404-639-2149; e-mail: jkm7@cdc.gov.

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Emerging Infectious Diseases
National Center for Infectious Diseases
Centers for Disease Control and Prevention
Atlanta, GA

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