Figure 1. This sequence shows HIV binding to the CD4 receptor, then to the CCR5 co-receptor, and finally having released its genetic material into a CD4+ T cellCD4+ T cell in four steps: (1) the CD4 receptor and CCR5 co-receptor on the cell surface; (2) HIV glycoprotein gp120 binds to CD4; (3) gp120 binds to CCR5 and releases gp41, which pierces the cell surface releasing the infective part of the virion into the cell; (4) the capsid (see HIV Structure & Function in the Glossary) enters the CD4+ T cell through the pore in the cell wall created in (3).

CCR5
CCR5 is a co-receptor on the surface of CD4+ T cells and some other cells that, during most of the course of HIV infection, is essential to entry of HIV into these cells. HIV attaches to both CD4 and CCR5 to achieve entry. (Some variants of HIV use a co-receptor called CXCR4 rather than CCR5; these variants almost always occur only late in untreated HIV infection; HIV transmitted from one person to another almost always uses the CCR5 co-receptor. However, some rare strains of HIV use both the CCR5 and the CXCR4 co-receptors.) Figure 1 (adapted from https://en.wikipedia.org/wiki/ccr5) shows (1) the CD4 receptor and the CCR5 co-receptor on the surface of a cell, (2) HIV binding to CD4, (3) then to CCR5, and finally (4) having released its genetic material into a CD4+ T cell through the pore in the cell wall created in (3). CCR5-tropic HIV is also called M-tropic because it can also infect macrophages and monocytes.

CCR5Δ32/Δ32
CCR5Δ32∕Δ32 indicates a mutation that deletes 32 nucleic acid base pairs from both parents’ copies of the gene that encodes the cellular co-receptor CCR5. The absence of these base pairs eliminates the ability of CCR5 to function on CD4+ T cells; the CCR5 co-receptor is needed by almost all strains of HIV to enter and infect these cells. Notably, the allogeneic immune-system transplants that resulted in a sterilizing cure of HIV infection in the Berlin Patient (Timothy Ray Brown) had this mutation in both strands of the DNA included in the transplant. Unfortunately, only about 10 – 15% of Caucasians have this mutation and almost no one else does, which makes this approach nearly useless for curing HIV infection in all infected persons, unless gene editing can make more instances of the mutation, which is one of the focuses of HIV cure research. Note that “Δ” is the upper-case Greek letter “delta” and stands for the deletion. Further, CCR5Δ32/Δ32 is associated with increased susceptibility to West Nile virus infection and a variety of encephalitis.

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CD4
CD4 is a receptor that is necessary, along with a co-receptor (CCR5 or CXCR4), to the attachment of HIV virions to CD4+ T cells. CD4 is one of hundreds of receptors known as clusters of differentiation that are found on the surfaces of various cell types and facilitate attachment of virions, chemicals, and other cells. CD8 is another. Note that, in addition to HIV-specific CD4+ T cells, CD4 is also found on other types of immune-system cells.

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CD4+ T Cells
CD4+ T cells are primary white blood cells of the immune system; they are also known as helper T cells. These cells act, for the most part, as the “directors” of the immune system; they signal to other immune-system cells how and when to fight infections. CD4+ T cells are preferentially infected by HIV, which causes its own genetic material to be converted from RNA to the corresponding DNA and to be integrated into the cells’ DNA. HIV-infected CD4+ T cells, when activated, produce copies of HIV instead of reproducing or conducting immune functions.

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CD4+ T cells can develop that specifically target parts of an infectious agent and such cells become activated in response to infection by that pathogen. After the infection is cleared or controlled, they can then become resting memory CD4+ T cells that lie in wait for future occurrences of the pathogen to which they then respond. Such resting memory CD4+ T cells are thought to constitute most of the latent reservoir of HIV. CD4+ T cells all have the CD4 receptor on their surfaces.

CD8
CD8 is a receptor that is necessary to the attachment of virions chemicals, and other cells to CD8+ T cells. CD8 is one of hundreds of receptors known as clusters of differentiation that are found on the surfaces of various cell types and facilitate attachment of virions, chemicals, and other cells. CD4 is another example. Note that, in addition to responding to HIV-specific CD4+ T cells, CD8+ T cells also respond to other CD4+ T cells and CD8 is found on other types of immune system cells.

CD8+ T Cells
CD8+ T cells are primary white blood cells of the immune system that kill infected or disabled cells as directed by CD4+ T cells. CD8+ T cells can be created that are specific to HIV. CD8+ T cells all have the CD8 receptor on their surfaces. These cells are also known as cytotoxic T lymphocytes (CTLs).

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Recent research suggests that harnessing the killing power of CD8+ T cells may be essential to both functional and sterilizing HIV cures (see the HIV Cure (Functional) and HIV Cure (Sterilizing)Glossary entries).

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Central Nervous System (CNS)
The central nervous system (CNS) consists of the brain and spinal cord. It is important for curing HIV for at least four reasons:

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1. it is a latent reservoir for HIV that is affected by chronic inflammation that begins very early in HIV infection;

2. it can only be reached by a small minority of HIV antiretroviral therapy (ART);

3. HIV’s gp120 glycoprotein impacts the function of neurons; and

4. since the brain is so very essential, there is concern among cure researchers that approaches other than shock and kill, such as latency silencing of reactivation entirely, will be necessary to achieve a cure in the CNS because of the seriously toxic effect reactivation is likely to have on CNS functioning.

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Also, several free-floating HIV proteins have been shown to enter neurons and have pathogenic effects.

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Clinical Trials
Clinical trials are the standard process for testing new medications, medical devices, and medical procedures in humans. They are typically preceded by studies done in nonhuman animals (sometimes called “Phase 0”) to weed out those that are not worth the effort and expense of clinical trials and to find the ones that are worth pursuing. Clinical trials have three phases, as follows (we use medication to represent all three categories below):

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• Phase I: A Phase I clinical trial involves a small number (usually not more than about 20) of healthy volunteers to test the safety of the medication and any side effects it may have. If the procedure is determined to be safe and to have only acceptable side effects, it may proceed to Phase II.

• Phase II: A Phase II clinical trial will usually involve several hundred volunteers. It continues to test for safety and side effects and also adds on determination of its effectiveness.

• Phase III: A Phase III clinical trial involves several thousand volunteers and is intended to confirm the effectiveness of the medication, monitor its side effects, compare it to commonly used drugs if there are any already, and continue to collect information to determine whether the drug is safe. Only after a successful Phase III study does a medication go before a panel of the U.S. Food and Drug Administration (FDA) or similar agency elsewhere in the world for approval for distribution.

There is also what is sometimes called “Phase IV”: a post-marketing phase, in which the medication or procedure is used in diverse populations and continues to be monitored for side effects.

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In some case, clinical trials have phases that are numbered with a Roman numeral followed by the letter “A” or “B” to indicate whether it is an early or late part of the phase, respectively. These letters are usually used in combinations of phase numbers to indicate that the clinical trial straddles two consecutive phases; an example of this might be a Phase IB/IIA trial.

Every clinical trial done in the United States is required to have a written trial protocol that describes at the least

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• a detailed plan of what is to be done,

• why it is being done,

• justification for it based on prior research,

• known and hypothetical risks and benefits,

• criteria for inclusion and exclusion of potential participants, anda schedule of what will be done (for example, physical exams, blood draws, and monitoring side effects).

It also must have an informed consent form (ICF) that explains the trial to the volunteers, informs them of the above facts in lay language, tells them they may withdraw from participation at any point without giving a reason for doing so, and requires their signed and witnessed consent to participation before they begin the trial’s procedures. Every clinical trial protocol and ICF is reviewed and approved by an independent Institutional Review Board and one or more of several government agencies (which one(s) depend on the nature of the trial) before it can begin recruiting volunteers. Additional resources for understanding clinical trials and the clinical-trial process (both for specialist and non-specialists; most providing access to the full text) can be found here.

Clinical trials done outside the United States are required to follow the same or a very similar rigorous plan and process.

Clonal Expansion
Clonal expansion is the production of numerous daughter cells with identical genomes resulting from a parent cell. Clonal expansion of HIV-infected  CD4+ T cells in circulating blood is thought by some researchers to be a significant contributor to the latent reservoir and so a barrier to HIV remission.

Co-Receptor
A co-receptor, in the context of HIV medicine (including cure research), is a chemical, such as CCR5 or CXCR4, attached to the surface of a cell such as a CD4+ T cell that facilitates attachment and entry along with a receptor, such as CD4, of an HIV virion into the cell.

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CXCR4
CXCR4 is a co-receptor on the surface of CD4+ T cells that, during late stages of untreated HIV infection, is essential to entry of HIV into these cells. (Some variants of HIV use a co-receptor named CCR5 rather than CXCR4; these variants almost always occur in all but the last part of the course of HIV infection; HIV transmitted from one person to another almost always uses the CCR5 co-receptor, though rare cases with the CXCR4 co-receptor do occur.) Further, some rare strains of HIV use both the CCR5 and the CXCR4 co-receptors. Also, unlike CCR5, CXCR4 is not a good candidate for gene editing because it occurs on several cell types other than CD4+ T cells and is essential to their function. CXCR4-tropic HIV is also called T-tropic because unlike CCR5-tropic virus it can infect CD4+ T cells but not macrophages and monocytes.

Defective Virion
A defective HIV virion is one containing an RNA genome that makes it incapable of viral replication. This results from the single-stranded nature of HIV’s RNA. All living organisms have linked double-stranded DNA making up the well-known double helix; the cross links in the helix provide a self-checking mechanism to prevent frequent mutations. Of course, some mutations do occur in living organisms, and they are one of the mechanisms that cause cancers and numerous other diseases, such as sickle-cell anemia and Huntington’s disease. However, the unlinked single strands of HIV’s RNA have no such self-checking mechanism, and mutations occur in them very frequently, as we shall see.

Let’s calculate how often a typical nucleic acid base is mutated each day in a person who is not on antiretroviral therapy (ART). Note that, despite the extremely high frequency of mutations, this has no chance at all of eliminating HIV from a human in a lifetime!

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1. The current best estimate for the overall mutation rate is one per 34 viral replication cycles.

2. Given that roughly 10 billion new virions are created each day, roughly 300 million of the new virions will have at least one mutation.

3. With about 9,750 nucleic acid bases in each strand or 19,500 across both strands, that’s an average of one mutation in each base position about 16,000 times a day!

Compared to a living organism’s mutation rate, this is absolutely staggering! It doesn’t require very many mutations in genes encoding critical proteins, such as reverse transcriptase or integrase (see the HIV Structure & Function Glossary entry), to render a virion incapable of infectivity, that is, make it defective. Even in persons on suppressive antiretroviral therapy (ART), the accumulation of mutations that make new virions defective is staggeringly common.

Dendritic Cell
A dendritic cell is one variety of antigen-presenting cell whose main function is presenting antigens found on external surfaces in the body to B cells or CD4+ T cells. They are found in the skin and other areas that are on the outside of the body, such as the nose, lungs, mouth, stomach, and intestines, and so in contact with the environment.

Diversity and Inclusiveness in Cure Research
It is no secret that HIV/AIDS is a pandemic disease, yet HIV-related research and cure research in particular tend very strongly to be concentrated in the developed world (particularly the United States, Canada, Western Europe, and Australia) plus a few relatively isolated outposts in Thailand and South Africa. There are issues of sex, gender, sexuality, age, race, economics, convenience, and researcher bias at the least that are responsible for this. Following are a few of the relevant facts and resources that make clear some of the issues and possible approaches to dealing with some of them; all of them are discussed in the Perspectives entry with the same header (in black).

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• It is clear from numerous studies that the immune system’s effectiveness decreases with increasing age. This has effects on how well the body can deal with HIV infection among many other types of assaults and it also probably impacts the effectiveness of approaches to HIV cure, though this is currently unknown.

• Similarly, the hormonal and other developmental changes that occur during adolescence and the legal issues involved in obtaining informed consent for research very often exclude adolescents from HIV research studies. A notable positive development in this area is in South Africa, the country with the highest prevalence of HIV+ youths and young adults.

• There are barriers to including women in cure research, at the least because current approaches to cure have unknown interactions with pregnancy, both on the mother and the fetus.

• Transgender health is a developing field, but, so far, virtually nothing is known about the impact of hormonal treatment and cure strategies.

Droplet Digital Polymerase Chain Reaction (ddPCR)
Droplet digital polymerase chain reaction (ddPCR) is a type of polymerase chain reaction that is characterized by the creation under digital control of tiny droplets of highly amplified DNA resulting from an initial single molecule that are digitally measured. This technique has greatly automated the PCR technology and markedly decreased its expense.

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Dual Antibody Use to Reduce the Latent Reservoir
Three studies published in late 2015 examine the use of so-called dual antibodies to achieve various aspects of reduction of the latent reservoir of HIV-infected CD4+ T cells, as follows:

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1. One study discusses the use of dual-affinity re-targeting (DART) molecules to bind to both the HIV Env protein and the CD3 receptor on infected cells and “recruit” CD8+ T cells to kill the bound cells.

2. Another study used DARTs to bind to the same protein and receptor as in item 1 above to direct CD8+ T cells to kill infected CD4+ T cells.

3. The third study used what it calls bispecific antibodies that bind to the Env epitope recognized by the broadly neutralizing antibody VRC07 and the CD3 receptor to reduce HIV DNA in most of the laboratory isolates of infected cells tested. A broadly neutralizing antibody is one that is effective against a large class of antigens. “VRC” abbreviates the Vaccine Research Center at NIH that was responsible for discovering and characterizing the antibody.

One potential concern about all three of these studies is the possibility that their targeting CD3 might cause general activation of T cells, since it is the receptor that occurs on all of them, which could be disastrous. Apparently the dual nature of the targeting caused only minor, transient effects of this sort.

Elite Controllers
Elite controllers are rare individuals living with HIV who maintain undetectable viral loads in the absence of—in most cases—any antiretroviral therapy. In about 2/3 of known cases, they possess immune-system mutations that appear to enhance immune-system recognition and removal of HIV. In some elite controllers, undetectable viral loads are found in the absence of protective genes, indicating that specific genes are neither necessary nor sufficient for elite control of HIV. However there is evidence that many elite controllers suffer from chronic systemic inflammation like other people living with HIV, so they are likely to suffer from its long-term effects.

Enzyme
An enzyme is an organic molecule, in most cases a protein or peptide but in a few cases an RNA (such as a ribozyme described in the Gene Editing Glossary entry), that acts as a catalyst: It facilitates a biochemical process without itself being modified, so it can be used again. Almost all proteins that are enzymes have names ending in “ase”.

Gene Editing
Gene editing is a cure strategy for modifying genetic information in cells, such as removing HIV proviral DNA from a person’s DNA or altering the CD4 receptor, CCR5 co-receptor, or anti-HIV restriction factors to make CD4+ T cells resistant to HIV infection. There are numerous experimental gene-editing techniques being investigated (many targeting the gene that encodes CCR5). We describe below only the most important one, namely, CRISPR. A recent mathematical modeling study of gene editing for HIV cure has shown that achieving positive results is possible only under a narrow range of conditions and that further improvements are likely necessary to improve outcomes.

CRISPR-based gene editing is a combination of two drugs, CRISPR (a DNA sequence originally derived from bacteria) and, usually, a Cas protein (CRISPR associated protein—most often Cas9), that is currently the most efficient, effective, and easy-to-use method for gene editing. A recent report discussed laboratory comparisons between older methods and uses of CRISPR/Cas9 to perform the same tasks and showed that there were erroneous results in a significant number of cases using the older methods.

In fact, Science, the most prominent U.S. scientific journal, declared CRISPR to be the “Breakthrough of the Year” for 2015 because of its very wide applicability and ease of use, and Nature, the most prominent British scientific journal, chose it as No. 1 among its ten most important breakthroughs of 2015. Further, in early 2016, it was reported by two research teams that CRISPR technology, one using Cas9 and another using a different protein, had been used to remove entire HIV proviral DNA from latently infected CD4+ T cells in vitro and this has more recently been reported in vivo. Problems remain, however, for generally translating this technology to use in vivo—in fact another recent study about it reported that CRISPR/Cas9 resulted in an immune response to the Cas9 protein as a substance foreign to the body (a pathogen).

See also the CRISPR Perspectives entry and the first item under the Gene Editing header in the Glossary.

Genome
A genome is the collection of all the genes in a living organism or virion.

Graft-versus-Host Disease (GVHD)
Graft-versus-host disease (GVHD), also called rejection, is a natural reaction by the body’s immune system to a graft or transplant that typically results in elimination of the graft or transplant unless immunosuppressive drugs, such as cyclosporine, are administered. The reaction is predominantly carried out by CD8+ T cells. Nevertheless, in the case of the Berlin Patient (Timothy Ray Brown), graft-versus-host disease may have played a significant essential role in destroying his original HIV-infected CD4+ T cells.

Gut-Associated Lymphoid Tissue (GALT)
Gut-associated lymphoid tissue (GALT) consists of immune cells lining the gut that are a critical component of the immune response to pathogens. It is usually severely depleted very early in the course of HIV infection. It is believed that the depletion is mostly irible.

HIV Cure (Functional)
This type of cure allows some infected cells to persist in the body of a person living with HIV but means that antiretroviral therapy is no longer necessary, at least for a long time. With this approach, the immune system should be able to handle the virus that is still in the body. Because such individuals would typically have very low levels of HIV, they would much be less likely to transmit HIV to others than most infected people but might themselves be vulnerable to reinfection with other strains of HIV than the one with which they are already infected. This type of cure is also commonly called HIV remission.

HIV Cure (Sterilizing)
This type of cure completely eliminates HIV from an infected person’s body, which would likely require activation and killing of all infected resting memory CD4+ T cells (and probably infected macrophages and other infected cells), plus eliminating or silencing other cells contained in latent reservoirs such as the central nervous system (CNS). Depending on the strategy used, such individuals might or might not be resistant to reinfection with HIV. This approach results in there being no HIV capable of viral replication left in the body, so the person would not be able to transmit HIV to others. However, proving that all HIV has been eliminated from a person’s body is impossible with current approaches, including in the case of the Berlin Patient (Timothy Ray Brown), though he has been HIV– for ten years.

HIV Genome
The nucleus of HIV (see the HIV Structure & Function Glossary entry) contains the two separate sngle strands of RNA tht make up HIV’s genetic material or genome. Each strand comprises nine genes and two long terminal repeats (LTR), one at each end, the right-hand one of which is crucial for beginning the production of proteins from the resulting proviral DNA. The overlapping of segments in the diagram corresponds to what are known as open reading frames. (Note that the open reading frames in the HIV genome are never directly transcribed and translated to proteins: The HIV genome must first be integrated into a host cell’s DNA as proviral DNA that is, in turn, transcribed and translated to proteins.) In all, each strand has roughly 9,750 bases (nucleic acids), though this varies somewhat with the faulty replication of HIV RNA (see both the Introduction Defective Virion entry and the Glossary Defective Virion entry).

HIV Structure & Function
The cutaway diagram in Figure 4 in the Glossary shows schematically the structure and components of an HIV virion. Note that Figure 1 in this Introduction shows the capsid about to be inserted into an about-to-be infected cell. The capsid contains everything necessary to insert the HIV genome (transcribed from RNA to DNA) into the cell’s DNA and to produce new virions. The components are described in detail in the Glossary entry with the same name.

HIV’s Uniqueness
HIV is unique among human pathogens in several respects, as follows (adapted in part from a slide created and provided for use by Nobel Prize winner Prof. David Baltimore, PhD, of CalTech):

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• It preferentially attacks CD4+ T cells, the “directors” of the adaptive immune system.

• It eludes control by antibodies.

• Sugars cover almost its entire accessible surface. The only notable exception is the CD4 binding site, but that site is deep inside the protein coat, where it can’t be reached by most antibodies.

• It employs a remarkable two-part attachment mechanism, using CCR5 or CXCR4 in addition to CD4. Entry only takes place after viral gp120 protein has bound to the CD4 site (see the CCR5Glossary entry). As a result, very few antiviral antibodies can neutralize HIV, and fewer still are both broad and potent. Broad refers to the range of variants of HIV that the antibody is acive against.

• It destroys the gut-associated lymphoid tissue (GALT) very early in infection altering the gut’s bacterial community.

• It also attacks the central nervous system (CNS) very early in infection.

All of these aspects of HIV’s uniqueness make it a much more difficult target for cure research than for almost all other pathogens.

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Immune System
The immune system is the body’s protection against disease. It consists of two major parts, the innate immune system and the adaptive immune system.

The innate immune system comprises three parts: biological barriers, natural killer (NK) cells, and killer-cell immunoglobulin-like receptors on the surfaces of natural killer cells, which are generally known by their abbreviation “KIR”. Biological barriers at the surface of the body may be effective in keeping out pathogens, such as foreign substances, bacteria, and viruses that the barriers recognize as different from the body. Pathogens that make it through the biological barriers may be recognized by KIR components that are specific to them. If a KIR component recognizes a pathogen, it activates natural killer cells.

The adaptive immune system comprises B cells, T cells, antibodies produced by B cells, and the human leukocyte antigen (HLA) complex, which consists of genes that code for body surface proteins that distinguish between self and non-self and cell-surface proteins, such as CD4 and CD8, that regulate the adaptive immune system in humans. T cells, in turn, are a large family of varieties, including at least CD4+ T cells, CD8+ T cells, and at least a half dozen other types. See the entries for the underlined cell types for descriptions of their roles in immunity.

Inflammation
Inflamed immune-system cells can signal other immune-system cells to reproduce or respond to a pathogen. The key white blood cell in inflammation is the macrophage. Macrophages can assemble within themselves specialized platforms named inflammasomes that produce the substances that promote inflammation. These platforms are assembled when needed and destroyed when they are no longer needed. This is usually helpful.

However, HIV infection, even in those whose virus is either suppressed naturally (elite controllers) or by antiretroviral therapy (ART), is known to cause chronic inflammation, which can lead to heart attacks, strokes, cancers, and other serious health conditions. Activated cells can also produce scarring (also called fibrosis) in lymph nodes, a critical part of the immune system. For most purposes chronic immune activation equals chronic inflammation (that is, every state of chronic inflammation leads to chronic immune activation and vice versa).

Latency Reversal
Latency reversal is fundamental to activating the bound HIV proviral DNA in resting CD4+ T cells in latent reservoirs in the body to make it susceptible to destruction in the approach to cure known as shock and kill. This is considered by many HIV cure researchers to be fundamental to curing HIV. The Glossary entry with the same name lists many types of substances and individual substances being tested as latency-reversal agents.

Latency Silencing
Latency silencing is a term used to describe an approach to completely stop reactivation of latently infected CD4+ T cells in latent reservoirs, thus making them incapable of producing further HIV virions. Latency silencing is essential to curing HIV in areas such as the central nervous system (CNS) where latency reversal is believed to have disastrous consequences. At least five distinct approaches are currently being explored, and NIH has a request for research-grant applications for another that is active as this is being written. Latency silencing is also called block and lock.

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Latent Reservoir
Latent reservoir is used in HIV cure research in two closely related senses, as follows:

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1. A latent HIV reservoir is a tissue in a person’s body that is reachable by HIV. In the context of HIV infection, it is a part of the body that, it is generally believed, is not affected by antiretroviral therapy (ART) as effectively, if at all, as in the blood. Latent reservoirs provide long-lived homes for HIV to reemerge from if therapy is stopped. Examples of confirmed and likely latent reservoirs include a subset of CD4+ T cells named resting memory CD4+ T cells, macrophages and monocytes (white blood cells), and parts of the intestines.  A recently reported study of HIV and macrophages performed in bone marrow-liver-thymus (BLT) mice showed that HIV persists in macrophages in vivo and thus they are very likely to be a component of the latent reservoir.

2. The latent HIV reservoir is the totality of the individual latent reservoirs of type (1). The size of the latent reservoir is estimated to be anywhere from 1 million to over 50 million HIV-infected resting memory CD4+ T cells by the methods listed in the Measuring the Latent Reservoir Glossary entry.

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A 2011 freely available survey article (Richman D “Introduction: challenges to finding a cure for HIV infection” Current Opinion in HIV and AIDS 6, January 2011, p. 1; it can be downloaded from the webpage http://journals.lww.com/co-hivandaids/toc/2011/01000 ) noted what we know about the latent reservoir as follows (lightly edited):

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1. Latently infected resting memory CD4+ T cells are the best characterized latent reservoir for HIV-1.

2. Less than 1 cell per million of resting CD4+ T cells from persons on potent antiretroviral therapy harbors replication-competent latent provirus.

3. Other drug-insensitive reservoirs, including brain, macrophages and hematopoietic stem cells, may also exist.

4. The genetic information in latent proviruses does not evolve—because it is produced by a clonal expansion of a single infected cell—which suggests there is no ongoing viral replication within the cells containing them. Discontinuation of antiretroviral therapy permits the rebound of viral replication originating from the latent reservoir.

5. Patients successfully treated with antiretroviral therapy for a decade or more exhibit no appreciable decrease in the size of the latent reservoir.

6. The persistence of the latent reservoir precludes its elimination by antiretroviral therapy for the lifetime of the patient.

7. Latency is likely established by numerous steps of HIV-1 replication, which potentially complicates eradication strategies.

It is generally accepted that the latent reservoir of at least HIV-infected resting memory CD4+ T cells containing proviral DNA and almost certainly other types of HIV-infected cells is established within days after infection.

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Lymph Node
A lymph node is a small organ containing immune-system cells named lymphocytes that filter lymph, which is a milky fluid similar in composition to blood plasma that contains fats (responsible for its color), B cells, and T cells; the latter include CD4+ T cells and CD8+ T cells. Prominent clusters of lymph nodes are found in the underarms, the groin, and the neck. See the Lymphatic System & Lymphoid Tissues Introduction and Glossary entries for more information about them.

Lymph Node Collagen Deposition (Fibrosis)
When cells die, they are sometimes replaced by scar tissue composed of collagen, which is a protein found in numerous tissues including bones. This is called fibrosis. When lymph nodes are inflamed by HIV viral replication they can lay down scar tissue. This can begin within days of HIV infection and may be largely complete within months after infection. Experts currently believe that when lymph nodes are scarred, it may be difficult to regain their ability to respond to HIV and other infections as effectively as before the deposition had occurred, causing lasting damage to the immune system that a cure may not be able to reverse.

Lymphatic System & Lymphoid Tissues
A lymphoid tissue is an individual component of the lymphatic system. The lymphatic system is made up of lymph nodes; local immune cells in many other lymphoid tissues, such as gut-associated lymphoid tissue (GALT), Peyer’s patches, the spleen, tonsils, and adenoids; and the lymphatic vessels that lead from lymphatic tissues toward the heart. The lymphatic system is essential to fighting infections.

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Measuring the Latent Reservoir
Measuring the latent HIV reservoir(s) is vital to determining the effectiveness of approaches to latency reversal. It can be used to determine the number of reactivated HIV-infected CD4+ T cells, in addition to its basic measurement role.

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It is estimated that the latent reservoir typically contains anywhere from about 1 million to over 50 million HIV-infected CD4+ T cells. The ultimate goal of measuring the latent reservoir is to count all and only replication-competent provirus, which no measurement tool is yet capable of doing. There are several approaches to measuring the number of HIV-infected CD4+ T cells in the latent reservoir, and more are being designed continually.

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The “gold standard” to which all other approaches are compared is the quantitative viral outgrowth assay (qVOA), which attempts to count replication-competent latent provirus. It is complex and expensive and has the added disadvantage of being very likely to significantly underestimate the actual size of the latent reservoir. However, some studies show a significant correlation between the results of qVOA and total HIV DNA.

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Natural Killer (NK) Cells
Natural killer (NK) cells are white blood cells responsible for killing infected cells and cancer cells. They are the most ancient component of the cellular immune system. They have long been thought to be purely “natural” in the sense that they are preprogrammed to respond to particular types of infected or disabled cells, unlike CD4+ T cells and CD8+ T cells, which must be trained to respond to their target pathogens and thus can have numerous distinct targets. However, recent evidence suggests that there are memory-like subsets of natural killer cells in mice and in nonhuman primate (NHP) models, such as rhesus macaques infected with SHIV. There is ongoing research into whether such memory-like natural killer cells may play a role in curing HIV infection.

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Post-Therapy Controllers
Post-therapy controllers are a small group of HIV+ individuals, so far mostly the VISCONTI (Viro-Immunologic Sustained CONtrol after Treatment Interruption) cohort in France, who started antiretroviral therapy (ART) within weeks of infection, stayed on therapy for an average of about four years, and then stopped therapy. Because there has been no large or lasting rebound of HIV, these individuals are able to stay off therapy for as long as 10 years. Unlike most elite controllers, these people mostly lack immune-system mutations that would make them less susceptible to ongoing virus replication. Natural killer (NK) cells are believed to be largely responsible for HIV control in this cohort.

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Receptor
A receptor, in the context of HIV cure research, is a chemical (such as CD4 and CD8). For a CD4+ T cell, the corresponding CD4 receptor facilitates attachment and entry along with a co-receptor, namely CCR5 or CXCR4, of an HIV virion into a CD4+ T cell.

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Remission
Remission is a term preferred by many researchers for HIV Cure (Functional). This is because functional cures, like cures for many types of cancers, may be short lived though they are likely to be repeatable at least for HIV.

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Retrovirus
A retrovirus is a virus, such as HIV, whose genetic material is RNA rather than DNA. Retroviruses are special in that they are able to integrate their RNA into the host DNA as proviral DNA, which enables the creation of new virions.

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RNA
RNA stands for ribonucleic acid. Unlike DNA, which exists only in the well-known double helix structure found in all living things or as single strands in some viruses, there are more than 30 forms of RNA with distinct functions. One form serves as the two unconnected strands of genetic material in HIV.

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Stakeholder Engagement
Stakeholder engagement refers to the involvement of essential people and organizations, including governments, foundations, research groups, companies, and especially individuals, in promoting understanding of HIV-related research, particularly clinical trials of both cure basic science and, potentially, curative processes; developing appropriate expectations; and sustaining involvement of persons in those trials. See also the Resource Guide entry with the same header.

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Thymus Gland
The thymus gland is located in the chest just below the neck. It is the origin of all T cells (including specifically CD4+ T cells and CD8+ T cells) all of which migrate to the bone marrow. The thymus gland typically shrinks to almost nothing during adolescence.

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Viral Load
HIV viral load measures the amount of HIV virions circulating in the blood. It is usually reported as copies of virus per milliliter of blood (abbreviated c/ml). It is important in HIV cure research because activating cells containing latent HIV from latent reservoirs increases viral load in a measurable way.

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Virion
A virion is a single complete virus particle that consists of an RNA or DNA core with proteins, such as enzymes, and often with an external envelope. It is the extracellular infective form of a virus.

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Women’s Involvement in Cure Research Studies
A recent open-access viewpoint article concerning women’s involvement in cure research suggests six ways to increase women’s involvement. Before summarizing the points in the article, we must point the reader to the Estradiol, Estrogen, Progesterone, and Estrogen Receptors Glossary entry, which makes clear several very important biological reasons for increasing women’s involvement. Current barriers and suggested ways to increase involvement are as follows:

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1. The possibility of pregnancy and its unknown or not clearly understood impact on HIV-related research of all kinds is a very frequent barrier, especially for treatment studies. Most sudy designs can be modified to reduce the impact of this barrier, if not eliminate it.

2. Researcher and clinic coordinator perceptions may impact recruitment of women.

3. Engagement of women stakeholders and improving the perceptions of women held by male stakeholders can increase women’s recruitment and retention in clinical trials.

4. Overcoming structural barriers, such as the lack of child care at research sites, and including women-focused community organizations in recruitment can improve involvement of women in studies.

5. Policy interventions in research funding can promote sex and gender equity.

6. The Gender, Race, and Clinical Experience (GRACE) study (a description of which can be downloaded from http:// online.liebertpub.com/doi/pdf/10.1089/apc.2013.0015) is an excellent example that specifically included recruitment of women and can serve as a model for other studies.

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Perspectives on HIV Cure Research

This section presents several of the most important trends, subject areas, and less than obvious reasons for HIV cure research, with references to items in the following two sections that are related to them.

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Chronic Inflammation
Chronic inflammation is a process that involves body-wide inflammation, which is very deleterious to overall health, and begins very early in the course of HIV infection. While antiretroviral therapy (ART) is highly effective at reducing or nearly stopping HIV replication, it lessens but does not eliminate chronic inflammation, which is implicated in premature aging, heart and circulatory problems, cognitive decline, and numerous other serious medical problems. All this makes it one of the most urgent reasons for cure research.

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CRISPR
CRISPR (clustered regularly interspaced short palindromic repeats) are segments of DNA in some bacteria and archaea that, combined with a protein such as Cas9 (CRISPR associate protein 9) or Cpf1 (centromere and promoter factor 1), constitutes a primitive immune system that is mostly effective against genetic sequences found in viruses, and plasmids (genetic structures in cells that reproduce independently of the DNA in the nucleus—see also the Organelle Glossary entry), two types of potentially injurious foreign material. The combination of CRISPR and either of the two proteins has been found to be the most effective and exact technique for editing DNA. It is particularly useful to HIV cure because it can be used to edit out the genes for the co-receptors CCR5 and CXCR4 and/or the entire HIV genome. In fact, CRISPR was recognized as the breakthrough of the year for 2015 by the journal Science and number one of the ten breakthroughs of 2015 by the journal Nature. It is fair to say that while there are researchers and companies still using several of the other techniques listed in the Gene Editing Glossary entry, CRISPR is very likely to eclipse them within the next few years—it already has a significant industry: as of January 2018 there were already at least three public companies and numerous others that were still private. Several research projects that used other gene-editing techniques have recently been repeated using CRISPR/Cas9, which has shown that some of them had incorrect conclusions.

Diversity and Inclusiveness in Cure Research
It is no secret that HIV/AIDS is a pandemic, yet HIV-related research and cure research in particular tend very strongly to be concentrated in the developed world (particularly the United States, Canada, Western Europe, and Australia) plus a few isolated outposts in Thailand and South Africa. There are issues of sex, gender, sexuality, age, race, economics, convenience, and researcher bias at the least that are responsible for this. The following are a few of the relevant facts and resources that make clear some of the issues and possible approaches to dealing with some of them:

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• It is clear from numerous studies that the immune system’s effectiveness decreases with increasing age (see, for example, for a very readable exposition, Chapter 2 “Things Fall Apart” in the book A Gawande Being Mortal pp. 25 – 54 Henry Holt and Co. New York 2014). This has effects on how well the body can deal with HIV infection among many other types of assaults and it also probably impacts the effectiveness of approaches to HIV cure, though this is currently unknown, in large part because most HIV research studies along with other HIV research have upper limits on the age of participants.

• Similarly, the hormonal and other developmental changes that occur during adolescence and the legal issues involved in obtaining informed consent for research very often exclude adolescents from HIV research studies. A notable positive development in this area is in South Africa, the country with the highest prevalence of HIV+ youths and young adults. There, for example, the Centre for the AIDS Program of Research in South Africa (CAPRISA) is enrolling not just adolescents but also children in research. An article about CAPRISA is available online here and CAPRISA’s website is here.

• There are barriers to including women in cure research, at the least because current approaches to cure have unknown interactions with pregnancy, both on the mother and the fetus.

• Initial research on the effects of female hormones on HIV cure research are described in the Estradiol, Estrogen, Progesterone, and Estrogen Receptors Glossary below and barriers to and some suggestions for increasing participation by women in cure research are discussed in the Women’s Involvement in Cure Research Studies Glossary entry.

• The AIDS Malignancy Consortium (https://web.emmes.com/study/amc/public/), a clinical-trials-sponsoring network of the National Cancer Institute that supports studies of cancers in HIV+ people and their treatment, has set up a laboratory in eastern Africa because of the expense and infeasibility of getting frozen samples to laboratories in the United States. This might conceivably be shared with cure research studies.

• Transgender health is a rapidly developing field, but so far almost nothing is known about the impact of combining hormonal treatment wth cure approaches. A wonderful recent book that explores transgender health issues is the book L Erickson-Schroth Trans Bodies, Trans Selves: A Resource for the Transgender Community Oxford Univ. Press New York 2014. While it devotes only about a dozen pages to HIV specifically, it also provides access to other relevant resources.

The Fundamental Problems
There are two fundamental problems in HIV cure research that are somewhat different from each other, as follows:

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• One is the need to very accurately measure the latent reservoir of HIV-infected cells so as to assess the effectiveness of both experimental approaches and likely the ultimate effect of clinical methods. Perhaps the most important example of both is the Mississippi Baby, who, when returned to medical care, was found to be infected. It was hypothesized that she may have had only a single HIV-infected cell at that time. This problem may have been largely solved early in 2018—see item 5 in the Measuring the Latent Reservoir Glossary entry.

• The other is the need to determine, once an effective approach to cure, be it sterilizing (see HIV Cure (Sterilizing)) or functional (see HIV Cure Functional)), has been developed what fraction of the infected population needs to be cured to achieve population-wide cure. This is akin to the question of achieving what’s known as herd immunity—the fraction of a population that needs to be vaccinated against a disease organism to protect the entire population from infection. This may end up being 100%, but it could be fewer.

Microbiome and Microbiota
A microbiotum (the singular of microbiota) is a collection of bacteria, archaea, fungi, and viruses that colonizes a particular part of a plant or animal’s body. The term microbiome was originally coined to refer to the genome of a microbiotum, but it is now increasingly used interchangeably with and more commonly than microbiota.

Our concern is with the human microbiomes; it is estimated that the genomes of all microbiomes in one person have roughly 200 times the genetic material of the human genome. Various parts of our skin have individual microbiomes, as do parts of the digestive system, the respiratory system, and other parts of the body. Further, microbiomes vary from person to person. Particular changes in microbiomes are associated with diseases. HIV alters human microbiomes, and they are involved in all aspects of HIV disease from prevention through treatment and cure. For example, very early in infection, the gut-associated lymphoid tissue (GALT) is severely depleted, and this changes the intestinal microbiome. In HIV cure research microbiomes are just beginning to be a subject of significant study. This is definitely an area to watch.

Remission (Functional Cure)
While the term “cure research” continues to be used somewhat indiscriminately, it is recognized by most researchers that a generally applicable approach to a sterilizing cure (see the HIV Cure (Sterilizing) Glossary entry) is extremely difficult to develop and is very unlikely to be developed any time soon. Thus, almost all cure researchers use the term remission or functional cure (see the HIV Cure (Functional) Glossary entry) in a similar sense to how it is used in cancer treatment instead. Remission would involve being entirely symptom free for at least several years—and, one hopes, preferably much longer—before requiring treatment to re-achieve remission.

Shock and Kill
Shock and kill has been the subject of intensive research in many laboratories, but it is increasingly recognized as likely to be insufficient to achieve a cure on its own. There are several reasons this is the case, but the most important ones are as follows:

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1. Until early 2018 we lacked the capacity to measure the latent reservoir accurately, so shock-and-kill studies may impact it, but we had no way to know to what degree they do; item 5 in the Measuring the Latent Reservoir Glossary entry describes a much more peomising approach. Nevertheless it is clear that current shock agents affect only a small fraction of the latent reservoir;

2. The sites of integration of the virus are very likely to affect the potency of a shock agent (latency-reversal agent), and they have off-target effects—we need more powerful and more specific agents and likely ones that also attack infected cells that have the same HIV integration site in their DNA (that is, cells that are the result of clonal expansion);

3. Any shock agent might induce infected cells to proliferate and expand, but they have not yet been paired with powerful enough kill drugs, such as, perhaps, immunotherapeutic agents, vaccines, or other types of drugs that induce cell death in esponse to HIV proteins.

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