Broadly neutralizing antibodies in HIV-1 treatment and prevention

Abstract

Antibodies that naturally develop in some individuals infected with human immunodeficiency virus 1 (HIV-1) and are capable of broadly neutralizing diverse strains of HIV-1 are useful for two applications: they can inform the rational design of vaccine immunogens, and they may be capable of preventing and treating HIV-1 infection when administered passively. A phase IIb study has been initiated with the experimental broadly neutralizing antibody (bnAb) VRC01, which has considerable breadth and potency (also referred to as a phase IIb HVTN 703/HPTN 081 and HVTN 704/HPTN 085 AMP efficacy trials) to evaluate its protective efficacy in individuals at risk of HIV acquisition. bnAbs prevent HIV-1 infection by selectively targeting vulnerable sites on the viral envelope (Env) protein that facilitates the entry of HIV. Although in very early stages, bnAbs capable of neutralizing a broad range of inter- and intraclade HIV-1 isolates have been demonstrated to have potential in treating patients either alone or in combination with antiretroviral drug therapy (cART); however, they are proposed to be advantageous over the latter as far as durability and side effects are concerned. Recent studies have indicated that combination therapy of potent bnAbs along with latency-reversing agents (LRAs) might also target latent reservoirs of HIV and kill them by recruiting effector cells, such as natural killer cells, thus confirming clinical progression. Possession of such qualities makes these new-generation potent bnAbs extremely valuable in effectively complementing the shortcomings of current ART drugs and improving the quality of life of infected individuals.

Introduction

Communicable diseases such as acquired immunodeficiency syndrome (AIDS), tuberculosis, malaria and neglected tropical diseases have been the scourge of millions of people in south-east Asia, home to a quarter of the world’s population. Despite our greater understanding of how human immunodeficiency virus 1 (HIV-1) manipulates host machinery in order to favor its own replication, leading to pathogenesis and transmission, a functional cure or protective vaccine remains elusive. Antiretroviral therapy (ART) has indeed significantly brought down the incidence of infection and improved the quality of life of those infected; however, ART possesses limited or no potential to interfere with the viral reservoir, and viral rebound has demonstrated association with treatment interruptions.^1,2

HIV-1 continues to be a major global health problem, with more than 35 million individuals (including about 1.8 million children) living with HIV infection in 2016 [as per the United Nations Program of HIV/AIDS (UNAIDS) data: http://www.unaids.org], with a global HIV prevalence of 0.8% among adults in 2015. Although the advent of highly active ART significantly has helped in reducing the incidence and improving the quality of life of HIV-infected individuals, both lack of adherence and poor access to ART are believed to be associated with the continued disease burden. It is estimated that approximately 46% of adults living with HIV had access to ART in 2015 (as per UNAIDS data). Despite its effectiveness, patients infected with HIV need to adhere to ART for life, as it does not eradicate infection. This is because ART drugs primarily target circulating viruses and are largely unable to access those that remain dormant in viral reservoirs as provirus. Hence, although current ART is essential in reduction of disease incidence, it alone is not sufficient in bringing comprehensive reduction in HIV incidence and functional cure. Recent discoveries of a novel class of highly potent and broadly neutralizing HIV-1-specific monoclonal antibodies, by way of single-memory B-cell (obtained from elite neutralizers) cloning technology, have provided new hope in treating patients. Some of these potent broadly neutralizing antibodies (bnAbs) have entered into clinical trials^4–8 and few more (either singly or in combinations) are currently being considered to enter into human clinical trials for assessing their efficacy and other clinical evaluations.⁸ They are currently being investigated for their possible role in compensating the shortcomings of ART drugs. In the present review, we highlight advancements in knowledge of the potential of bnAb-mediated protection and treatment.

What are bnAbs and why are they considered important in disease prevention?

Neutralizing antibodies are specific types of antibodies that have the ability to act on the specific sites on microorganisms such as viruses that aid in establishing an infection. In bacterial infections, antibodies are known for neutralizing toxins, enabling opsonization and facilitating complement-mediated lysis; in viral infections, antibodies interrupt transmission primarily in two ways: by blocking virus entry into the uninfected target cells (by way of disengaging virus–receptor interaction) and via antibody-dependent cell-mediated cytotoxicity (ADCC). While neutralizing antibodies are generally effective in tackling type-specific microbes with limited or no genetic variations, for complex viruses such as HIV-1 and influenza (which display considerable variations in their genotypes), ‘very special’ antibodies capable of efficiently tackling the breadth of these genetic variations are essential. These ‘special antibodies’ are referred to as bnAbs. Since neutralizing antibodies exclusively target the viral surface protein that enables virus entry (the first step in establishing infection), bnAbs are being considered as the ‘blueprint’ for preventive vaccine design, especially for HIV-1 whose genotype varies considerably.^9,10 During the course of natural infection, while individuals infected with HIV-1 typically develop type-specific cross-neutralizing antibodies,¹¹ only about 1% of them are found to develop broad and very potent neutralizing antibodies (bnAbs)¹² that can neutralize a wide range of genetically diverse HIV-1 subtypes. Although the precise mechanism by which this rare fraction of chronically infected individuals develop bnAbs in the natural course of infection has remained elusive, recent structural, virological and immunological studies have provided strong evidence of how virus–antibody coevolutions in the natural disease course are synergistically associated with sequential development of potent and broad neutralizing antibodies via somatic hypermutations and maturation of antibody genes.¹³ The extent of suppression of viremia in chronically infected patients by bnAbs is not very clear; nonetheless, because of their ability to directly act on the viral envelope (Env) protein and thus prevent HIV-1 entry, bnAbs hold potential in treating infected patients (through ‘passive’ immunizations) in addition to ART currently given.¹⁴ Additionally, since bnAbs can also latch onto the viral Env expressed on the infected cells, they could potentially facilitate fragment crystallizable (Fc)-mediated clearance.¹⁵ Although initiation of serum-based immune therapy for treating infectious diseases goes back to the late 1800s,¹⁶ as well as the considerable success of monoclonal antibody (mAb)-based cancer therapy in recent times,¹⁷ there has been a huge gap in its application in treating infectious diseases.¹⁸ In the next section, evidence is highlighted regarding advances made toward understanding the therapeutic potential and nature of applications that bnAbs can offer.

Evidence of bnAb-mediated protection and treatment against HIV-1 and simian immunodeficiency virus (SIV)

Antibodies have been historically used in prophylaxis and treatment of several bacterial and virus-induced infectious diseases,¹⁹ and have had success in diverse infections [such as respiratory infection-causing bacteria like Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae, tetanus, Clostridium botulinum, herpesvirus (cytomagalovirus and varicella zoster virus), enterovirus infection in adults, and vaccinia].¹⁹ In fact, the first licensed monoclonal antibody (mAb), palivizumab (licensed in the USA in 1998), was found to prevent infection in infants who were at risk of contracting respiratory syncytial virus infection.²⁰ In early studies with passive immunotherapies, patient sera provided evidence of improved clinical outcomes associated with reduction in plasma viral ribonucleic acid load and delayed appearance of AIDS-defining illnesses.^21,22 In contrast to the polyclonal serum samples, due to their homogenous composition, mAbs have higher specific activity than that of serum antibodies and hence provide greater therapeutic efficacy.³ The first-generation bnAbs to HIV-1 such as b12, 2F5, 4E10, 2G12 and KD-247 were used to assess the extent to which they could reduce viremia in infected patients by passive infusion.^23–28 Although a transient decrease in plasma viral load and delayed viral rebound was observed post mAb transfusion, frequent virus escape due to emergence of resistance mutations against these mAbs was observed.^25,29,30 Isolation of potent bnAbs from elite neutralizers has not only highlighted key vulnerable sites on the viral (HIV-1) Env that the virus uses to evolve in the natural course of infection, but also makes them amenable to effective use in designing improved immunogens via reverse-vaccinology approaches³¹ and treatment through passive immunization.^32,33 Some of these potent bnAbs such as VRC01, 3BNC117 and PGT121 have shown promising results in animal models, as well as in humans, and have shown modest and transient suppression of viremia,¹⁴ thus opening a very interesting and important avenue of exploring how bnAbs can be successfully used as therapeutic agents to complement ART in comprehensive treatment of HIV-infected patients.

Significance of baseline viremia on the degree of efficacy of passively infused bnAbs

When passively administered to patients, can bnAbs be more efficacious when circulating viral load is low? Recently, two very potent and broad mAbs that target CD4 binding sites (CD4bs) on the viral Env, 3BNC117 and VRC01, were assessed for their degree of efficacy when administered to HIV-infected patients with low viremia. In a phase II open-label trial, when administered to patients with analytical ART treatment interruption (ATI), 3BNC117 mAb was found to be associated with a significant delay in viral rebound compared with what was found during ART alone.⁵ Comparable evidence of delayed viral rebound due to virus escape was also found in ATI patients infused with another CD4bs-targeting bnAb, VRC01 (National Institutes of Health [NIH] 15-I-0140 and AIDS Clinical Trials Group [ACTG] A5340 trials);⁷ however, unlike in the case of the 3BNC117 efficacy trial, profound delay was not observed. Taken together, both these studies highlighted how delayed viral rebound during monotherapy is dependent on low viral load set point in ART-experienced patients who do not harbor pre-existing viruses resistant to the bnAb used for therapy. Nonetheless, combination therapy with a cocktail of bnAbs with distinct specificities on the viral Env holds better promise than monotherapy with regard to minimizing viral escape and rebound.^5,7,34

Ability of bnAbs to interfere with latent viral reservoir and clearance

While ART has been shown to significantly improve the quality of life of infected individuals, it largely lacks the ability to directly modulate latently infected cells and hence is, in general, ineffective in curing persistent HIV infection.^35,36 Although it has not yet been ascertained whether bnAbs either alone or in combination can significantly limit the size and stability of the latent virus reservoirs, a few studies have provided evidence that bnAbs could possibly act on latently infected cells. For example, bnAbs like VRC01 and PGT121 have been demonstrated in vitro to block HIV-1 replication in activated cells that form the viral reservoir.¹⁴ Moreover, while 3BNC117 was found to possess the potential to kill latently infected cells, PGT121 administered passively to nonhuman primates has demonstrated association with significantly depleted proviral DNA.³⁷ Because of their ability to directly bind to viral Env expressed on infected cells, potent bnAbs are thought to be able to latch onto HIV-1 Env expressed on activated cells harboring proviral DNA, which will have dual implications: enable killing of latently infected cells by recruitment of natural killer (NK) cells¹⁴ and prevent further forming of latently infected cells by neutralizing HIV-1 released from activated reservoirs. Bruel and colleagues³⁸ recently demonstrated Fc-region-mediated killing of HIV-infected lymphocytes via ADCC activity and engagement of NK cells, mediated by some of the bnAbs that have the ability to limit cell-to-cell HIV-1 transmission.¹ While a number of approaches have been proposed toward reactivation of latently infected cells by applying agents that act on cellular transcription factors and host cellular activation signals such as protein kinase-C (PKC), toll-like receptor (TLR) and Mitogen-activated protein kinases (MAPK) agonists, latency-reversing agents (LRAs), and checkpoint blockers, the use of LRAs along with bnAbs for greater virus suppression has gained special interest.^39,40 Although no LRA has been reported that can very effectively act on cells latently infected with HIV-1, it has been proposed^41,42 that LRAs such as histone deacetylase inhibitors (HDACi) and TLR antagonists (e.g. vorinostat, romidepsin and panobinostat), TLR7 agonists (e.g. GS-9620) and other viral inducers such as I-BET151 and anti-CTLA-4 may work to limit the size and functionality of the latently infected cells when administered with potent bnAbs. Indeed, in experimental humanized mice, in conjunction with different LRAs described above, bnAbs 3BNC117, 10-1074 and PG16 led to beyond-detectable cell-associated proviral load.^14,43 Taken together, along with effective LRAs and ART, bnAbs could significantly help to limit the frequency of plasma viral rebound by reducing persistent HIV reservoirs.^44,45

bnAbs that block cell-associated HIV-1 transmission act as a ‘double-edged sword’

Typically, the prevention of cellular entry by bnAbs is tested on cell-free viruses; however, cell–cell spread is considered to be a more robust mechanism of HIV-1 spread and transmission.¹ Therefore, identifying bnAbs capable of also inhibiting cell–cell transmission would theoretically provide better clinical outcomes during therapy. bnAbs such as those targeting CD4bs (NIH45-46 and 3BNC60) and V3-glycan (10-1074 and PGT121) have been reported possessing properties to neutralize both cell-free and cell-associated viruses¹ by acting on the virological synapse that forms when an infected cell contacts an uninfected permissive cell (such as a CD4⁺ T cell).^46,47 bnAbs with the ability to interfere with both cell-free and cell-associated virus transmissions are generally believed to provide better treatment efficacy over the ones that are only effective in neutralizing cell-free viruses. This was perhaps the reason why bnAbs such as 10E8 and 3BC16 that effectively neutralize the majority of cell-free viruses were found to be ineffective in suppressing viremia in vivo,^1,34 while dually effective 10-1074 and NIH 45-46 mAbs were effective in modestly suppressing viremia in vivo.³⁴ Although the impact of neutralizing antibodies on the macrophage–T-cell-mediated virological synapse have been documented,⁴⁸ how neutralizing antibodies act on dendritic cell-mediated cell–cell transmission remains elusive (dendritic cells play a very important role in initial infection and viral transfer).⁴⁹ Taken together, potent bnAbs capable of preventing cell-free and cell–cell virus transmission are likely to exert better clinical outcomes when used in passive therapy and would be effective in tissues that support HIV infection such as gut-associated lymphoid tissue and lymph nodes.^1,50–52

Modification of bnAb arms and implications for improving therapeutic efficacy

The discovery of potent bnAbs has also paved the way for their further modification in order to enhance their utility in vivo for better clinical outcomes when used in therapy. Modifications that prolong half-life, and increase potency, Fc receptor (FcR) binding and polyfunctionality are thought to circumvent the shortcomings of bnAbs when used as whole molecule, and provide durable and long-lasting protection during passive immunotherapy.^53,54 A number of studies have demonstrated how modifications of the neonatal FcR (FcRn) can help maintain immunoglobulin (IgG) homeostasis and prolong the serum half-life of IgG in humans.⁵⁵ Such modifications are also believed to enhance mucosal localization, efficient immune protection, and durability of bnAbs during passive immunotherapy compared with their wild-type counterparts.⁵⁶ Selective modification of fragment antigen-binding regions (Fabs) of bnAbs has also been demonstrated to considerably enhance potency and breadth of bnAbs. In recent times, bispecific and trispecific bnAbs have gained special attention for their ability to comprehensively arrest virus infection by synchronously latching onto distinct viral and cellular targets that support HIV-1 entry. Very recently, considerable breath and potency of single antibody molecules engineered to possess three distinct specificities to HIV-1 Env have been reported.⁵⁷ The investigators demonstrated that the combination of three bnAbs with specificities to CD4bs (VRC01, N6), V1V2 (PGDM1400) and membrane proximal external region (MPER) (10E8v4) into one single molecule conferred very potent and broad neutralization in a nonhuman primate model, beyond what each of these individual bnAbs could achieve on their own. In addition, recent reports highlighted the potential of bispecific antibodies with dual specificities to provide considerable potency and neutralization breadth.^54,58–60 In one study, the antibodies were engineered such that one arm of the Fab binds to the cellular receptor (e.g. CD4 or CCR5) and the other Fab arm binds to epitopes on viral Env that are targeted by bnAbs. The second group⁵⁹ constructed a bispecific antibody by putting together the Fabs of two different potent bnAbs with distinct epitope specificities on viral gp120 with a goal to crosslink gp120 domains within a trimer spike. Both the approaches were reported to provide far better virus neutralization activity via their synergistic action over parent antibodies, which suggested the additive effect exerted due to antibody engineering. Antibody engineering has also been suggested as a means to effectively target the latently infected viral reservoir.¹⁴ The idea is to engineer antibodies in such a way that they can dually act on the virus as well as the activated cells that harbor HIV-1 in a dormant state. Two classes of such antibody molecules have been investigated: bispecific T-cell engagers and dual-affinity retargeting molecules. Because latently infected cells such as resting CD4⁺ T cells are not accessed by ART or most antibodies to HIV-1, upon activation, they are believed to express viral Env on the cell surface, enabling bnAbs to latch onto them and facilitate the recruitment of cytotoxic effector cells to kill by ADCC.⁶¹ Rational engineering of bnAbs makes them an attractive prospect, by providing additional effector functions beyond virus neutralization and prolonging virus suppression and eradication of viral reservoirs.

Summary

With the advent of single-memory B-cell cloning technology, scientists have been able to discover a number of very potent and broadly neutralizing human monoclonal antibodies from infected individuals. These bnAbs not only are aiding the design of improved immunogens but could also be useful in treating individuals infected with HIV-1. In sharp contrast to the drug regimens that are currently being used as ART, bnAbs have the potential to access latently infected activated T cells in addition to cell-free viruses in the circulation, with help from LRAs. Clinical trials with new-generation promising potent bnAbs are underway to assess their effectiveness in comprehensive treatment, along with their safety, durability and tolerability properties, which should be able to significantly reduce disease burden. Moreover, for effective control of HIV-1 replication, which shows extraordinary diversity in the clinical setting, combination therapy using a cocktail of bnAbs with multiple specificities would concomitantly target distinct and discontinuous epitopes on the viral Env, thereby significantly impeding viral escape and rebound.^34,37,62 Finally, while discovery of promising bnAbs to HIV-1 with potential in treating patients are encouraging, there are two areas that need particular attention prior to taking promising bnAbs for clinical development. First, the bnAbs may not necessarily provide comparable breadth and potency toward neutralizing HIV-1 in vivo to that observed under in vitro conditions, as demonstrated recently.^63,64 Hence, it will be important to assess the extent (both potency and breadth) to which the promising bnAbs (either as unmodified or engineered forms) neutralize HIV-1 grown in physiologically relevant cells, for example, peripheral blood mononuclear cells (or CD4⁺ T cells). In addition, it is important to assess the fitness cost of the bnAbs associated with emergence of escape variants of HIV-1.^4,64,65 For example, some bnAbs, if given as monotherapy, may select for early escape variants over others that would impact on treatment progress. Understanding such phenomena will help to choose the bnAb combinations that would overcome this problem and provide better treatment outcome. Secondly, it is important to periodically monitor circulating HIV-1 strains across populations and geographical boundaries to assess the breadth, potency and suitability of promising bnAbs.^66,67 Typically, viral Envs routinely used to assess the breadth and potency of bnAbs represent viruses which were either isolated in the past or which do not necessarily represent what is currently circulating. This may impact on the bnAb efficacy in clinical trials. In summary, advances made in the discovery and effective antiviral application of bnAbs in HIV-infected patients will provide guidance in developing mAb-based therapies for other emerging and re-emerging pathogens.