INTRODUCTION

HIV/AIDS remains a global health crisis, despite significant progress in antiretroviral therapy (ART). Although ART effectively suppresses viral replication in the bloodstream, it cannot completely eradicate the virus due to the establishment of viral reservoirs within highly divers tissue compartments and cell subsets. These reservoirs, which include a pool of long-lived latently infected cells, serve as a major obstacle to achieving an HIV cure. Therefore, developing strategies to reach, reactivate, and eliminate HIV persistence in tissues is a major goal in the HIV field.

Antiretroviral treatment interruption (ATI) studies have shown that viral rebound can arise from many different tissue compartments [1–3]. As small numbers of cells with intact HIV provirus can cause viral rebound after ATI [4▪], identification and characterization of the HIV cellular reservoir in tissue is imperative for developing elimination strategies. Considering that cell phenotypes, metabolic requirements and activation states differ between tissue compartments, efficient delivery of tailored therapeutic agents to tissue reservoirs represents a major challenge in targeting HIV persistence. Here, we focus on the most recent advances in literature of the HIV tissue-reservoir and emphasize strategies to reach relevant tissue compartments. General induction of cytotoxic cellular responses and reversal of cellular exhaustion features surpass the scope of this review. We argue that future research should aim for a multifaceted and tissue-specific approach that considers the early dissemination in multiple body compartments. 

Box 1:

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TISSUE-RESERVOIR ESTABLISHMENT

HIV persisting reservoirs are rapidly established early after infection [5▪▪,6,7], as well as at the time of ART initiation [8]. Translationally inactive and genetically intact proviruses can be detected in human blood and lymph nodes (LN) even before peak viremia [5▪▪]. Mucosal portals of entry such as the anogenital and urogenital tracts potentially represent the first site of HIV replication and reservoir establishment, including tissue-resident memory T cells (TRM) [9] and resident myeloid cells [10]. Subsequently, rapid HIV dissemination from these sites to CD4+ T-cell rich lymphoid tissues (LT) and the gastrointestinal (GI) tract implies formation of HIV persistence throughout the body (Fig. 1). CD4+ T cells from LT have been shown to rapidly produce CCL2 as an innate immune response during early stages of HIV infection, attracting infected CCR2/CCR5+ CD4+ central memory T cells (TCM) towards the tissue, likely expanding the reservoir [11]. However, the impact of ART and acute infection resolution on long-term memory persistence in the body remains largely undefined.

FIGURE 1:

HIV reservoir establishment throughout the body. HIV viral particles penetrate the mucosal epithelium and infect CCR5+CD4+ T cells from the submucosa in a process facilitated by mucosal myeloid cells. Local cellular infection and viral spread result in an inflammatory micro-environment. In this compartment, infected CD4+ circulatory memory T cells (TCIRC), CD4+ resident memory T cells (TRM), and macrophages (Mϕ) may already establish a local reservoir. Other infected TCIRC and myeloid cells disseminate HIV towards the draining lymph nodes (LN), where infection expands and, through blood, spreads to multiple compartments. The immune response in the LN follicle may already establish a TCIRC/TRM reservoir in the T-cell zone, as well as a CD4+ T follicular helper reservoir (Tfh) in the B-cell zone. Simultaneously, systemic high viremia and inflammation attracts an immune cellular response to tissues to contribute to viral clearance, including antigen-specific and bystander HIV-infected CXCR3+CD4+ T cells and peripheral blood monocytes, among others. Thus, infected T cells and monocytes reach multiple tissues, ultimately establishing a wide variety of HIV reservoirs within other lymphoid tissues, the central nervous system, the gastrointestinal tract, organs and peripheral tissues. All these compartments contain reservoir cells with specific and tissue-unique features.

HIV-infected peripheral blood CD4+ T cells are enriched in CXCR3 expression during early stage infection [5▪▪], indicating an effector Th1 response migrating towards inflamed tissues. Whereas most migrating CD4+ T cells are short-lived, part will develop into long-lived cells that remain at tissue locations [12▪]. Thus, an immune response, which likely includes antigen-specific HIV-infected cells, initially generated in LT and distributed to multiple inflamed anatomical compartments during acute viremia, may eventually establish a protective TRM response in infected compartments, potentially contributing to long-term HIV persistence [9,12▪,13▪]. Considering the antigen-specific CD4+ T cell response generated in macaques that were intravenously exposed to live-attenuated SHIV viruses, the consistent detection of Gag-specific CD4+ T cells in tissues such as the vaginal mucosa [14], may provide evidence for dissemination and persistence of long-term memory. Moreover, a limited number of HIV variants infects a large pool of clonotypically distinct T cells in both blood and LN [5▪▪], suggesting that activated CD4+ T cells specific to common antigens may be early targets for HIV and potential long-lived reservoirs [5▪▪]. This is exemplified in a recent case-report [15▪▪], which showed HIV reactivation and spread driven by a CD4+ T cell response to a co-pathogen in the brain.

Thus, besides HIV infection per se, other inflammatory processes may dictate the rate of HIV reactivation, dissemination, and redistribution throughout the body, as they affect tissue resident cell responses and maintenance. Indeed, a recent mathematical model implies that the contribution of cellular proliferation and differentiation exceeds the rate of HIV clearance in memory subsets from blood [16▪]. Since clonal expansion has been shown as a major mechanism of persistence [17–20], both homeostatic and antigen-driven proliferation will likely contribute to this phenomenon in tissues. New approaches allowing simultaneous determination of HIV intactness and cell phenotype demonstrate that CD4+ T cells harboring intact, inducible, and translation-competent genomes during ART are enriched for α4β1 expression [21▪▪], which supports the idea of redistribution of clonally expanded cells into inflamed tissues.

HIV PERSISTENCE IN TISSUE COMPARTMENTS

Identifying the cellular hallmarks of the HIV reservoir is imperative for developing new strategies to achieve reservoir elimination. In human peripheral blood, recent studies show that HIV-infected memory CD4+ T cells contain gene expression patterns that promote cell survival, proliferation, and HIV silencing [22▪], and these cells express higher levels of co-inhibitory receptors and ligands compared to uninfected cells [23▪]. Additionally, multiple cellular subsets besides well established CD4+ TCM, effector memory T cells (TEM) [24] and circulating follicular helper T cells (Tfh) [25], such as mucosal associated invariant T cells and Th2, may harbor integrated proviral DNA under ART [26▪]. Together with potential resistance of CD4+ T cell reservoir cells to immune-mediated killing [22▪,27], efforts to eliminate the reservoir may thus be further complicated by cellular and epigenetic heterogeneity of the reservoir between and within individuals [26▪].

It remains to be established whether tissue-resident HIV reservoirs also exhibit multiple levels of cellular, epigenetic and individual heterogeneity. TRM cells in barrier sites show vast functional and phenotypical adaptations to the specific site they reside in, contributing to their cellular heterogeneity [13▪]. Indeed, CD4+ T cells from skin and jejunum showed negligible overlap with other sites [13▪], highlighting the need for evaluating multiple tissues when evaluating viral persistence in TRM. In addition to T cells, HIV-infected monocytes in peripheral blood may reseed the tissue reservoir by recruitment of monocyte-derived macrophages to inflamed tissues, contributing to the early establishment of the reservoir in tissues such as the lung or liver [28,29▪▪]. Macrophages can be long-lived resident immune cells and highly diverse based on their origin and tissue conditions [30]. Considering the participation and compartmentalization of myeloid cells in viral persistence, these cells represent an additional challenge to viral persistence elimination [31▪▪].

New approaches to timely and sensitively detect the functional cellular HIV reservoir in blood may be of help to predict the size of the viral reservoir in peripheral tissues [32]. This may potentially be achieved by considering tissue-homing properties of cells that transiently appear in peripheral blood, yet the challenge remains based on nonrecirculating resident reservoir cells. In the following sections, we discuss tissue-specific characteristics of the HIV reservoir and emerging efforts to reach and target these cells (summarized in Fig. 2).

FIGURE 2:

Strategies to reach the HIV reservoir in tissue compartments. Multiple therapies in the central text are proposed that can be directed towards various anatomic compartments, potentially aided by homing molecules. The anatomic compartments include the central nervous system (in pink; top), gastrointestinal tract (in yellow; bottom), lymphoid tissue (in blue; left) or other tissues (in green; right). Specific interventions discussed in the review for each of these compartments are summarized in the drawing. In other tissues, a box entitled “other reservoir-reduction strategies” highlights recent interventions that can be systemically applied towards reservoir elimination in multiple tissues (including other colored compartments).

LYMPHOID TISSUE

LT harbors most of the HIV reservoir. Therefore, strategies to enhance cellular trafficking towards these areas or block establishment of LT reservoirs may provide steps towards reservoir elimination. Persistence of HIV has been shown in LN of acute HIV infected patients despite early ART and plasma viral suppression [33]. This persistence is likely achieved through proliferation of infected cells [16▪,34▪].

LNs are highly structured secondary lymphoid organs organized in B cell areas, which include Tfh-rich germinal centers (GC), and T cell areas harboring TRM, amongst other cell types [12▪]. Whereas the pool of infected cells in LN during chronic infection has been shown to be highly heterogeneous, the majority of HIV infected cells within the LN are CD4+ Tfh cells [23▪,26▪], and higher levels of HIV RNA correlate with higher proportions of germinal center CXCR3+ Tfh cells [33]. However, cells carrying intact provirus were equally distributed between Tfh and CD4+ TRM, expressing higher levels of CD44, IL-21R, CD127, and CD28, which suggests enhanced cell survival [23▪]. Thus, at least two different CD4+ lymphocyte compartments need to be targeted for viral persistence in LT; one located in the B cell follicles (BCF) and another one located in the T cell zone.

Expression of CXCR5 and attraction by CXCL13 are required for B cells and Tfh cells to enter the follicular GC in the B cell zone [35]. Antigen-specific Tfh cells make up ∼7% of the resident CD4+ T cell population in LT [36] and comprise a substantial heterogeneous pool of cells, as shown by their differentiation into Tfh1, Tfh2, and Tfh17 cells [12▪]. Promoting infiltration of effector CD8+ T and NK cells through the expression of CXCR5 into the follicles is a major strategy towards elimination of HIV persistence. CXCR5+CD8+ T cells have been shown to control viral replication within GC, especially in spontaneous controllers [35]. Moreover, adoptive transfer of CXCR5+ Chimeric antigen receptor T cells enhanced LN migration and reduced viral load in SIV-infected macaques, although these cells did not reach all tissue sites and were undetectable after 2–4 weeks [37]. Similar to CD8+ lymphocytes, CXCR5+ NK cells present within BCF of SIV-infected macaques show enhanced functionality compared to CXCR5- NK cells and negatively correlate with viral RNA [38]. Several treatments in nonhuman primate models have been reported to enhance CXCR5 expression, LN homing, and functionality of NK cells, such as B-cell derived interleukin (IL)-6 [39], IL-12/IL-15 [38] and IL-21/interferon (IFN)a [40].

Alternative strategies involve targeting factors that promote reservoir establishment and maintenance of infected CD4+ T cells. Transforming growth factor (TGF)-β has been shown to promote HIV infection of resting and activated memory CD4+ T cells by induction of CCR5 expression in vitro and possibly contribute to reservoir establishment by enhancing expression of LN-homing receptor CCR7 on these cells [41]. Furthermore, in vivo blocking of the TGF-β type-1 receptor by galunisertib in SIV-infected macaques on ART simultaneously reactivated the HIV reservoir and increased anti-SIV antibody and T cell responses [42]. Thus, blocking TGF-β may contribute to limiting reservoir establishment by actively reactivating infected cells, acting as a latency reversal agent (LRA). On the other hand, in humans, TGF-β stimulation in vitro increased CD8 frequencies in follicular areas of the LN [43]. Thus, considering the different needs for modulating CD4+ and CD8+ T cells, as HIV targets and immune effectors respectively, these type of therapies may need to be individually addressed for each particular subset.

Last, in situ imaging of protein and nucleic acid present within LN of SIV-infected macaques has shown that approximately 30% of infected cells were macrophages [44▪▪]. B cells were shown to attract these macrophages through IL-10 secretion, polarizing them to IL-10-producing M2 macrophages and possibly creating a suppressed environment for the SIV reservoir. Indeed, SIV-infected macaques show presence of IL-10 in the BCF and neutralization of soluble IL-10 impaired survival of the CD4+ T cell reservoir [45▪]. Together, the HIV reservoir in LN shows a wide array of cell types which needs to be targeted considering its heterogeneity and location.

GASTROINTESTINAL TRACT

The gut mucosa and gut-associated lymphoid tissue (GALT) represent a large part of the HIV reservoir [46], containing more activated CD4+ T cells and tissue-resident HIV reservoirs than any other human tissue site [47]. An additional complicating factor for this compartment is the differing immune composition and HIV burden between anatomical sites [48,49].

Th17 cells are highly susceptible to HIV infection and their loss across the GI tract contributes to a disruption of intestinal homeostasis and increased inflammation [50]. Failure to restore CCR6+ Th17 cell levels in the GI tract after HIV infection may be due to out-competition by CCR6+ Th22 cells for their shared migration towards CCL20 chemokine secreted by duodenal enterocytes [51]. Therefore, interference in this migration pathway may provide new strategies to enhance Th17 restoration into the GI tract. Still, the fraction of Th17 cells that survive account for the majority of the HIV reservoir in the rectum [52]. The transcription factor RORC2 was recently described as a host cofactor in HIV replication in Th17 cells, providing a potential target for HIV reservoir elimination [50]. Further, mucosal vaccination may offer new venues to directly impact cells in this compartment, since restoration of the Th17/Treg ratio in rectal biopsies of macaques was achieved by vaccination with an oral polio vaccine modified to secrete HIV-antigens [53].

Additional reservoirs to consider in the GI tract are Tregs and macrophages. In macaques, CD101+CD4+ T cells, the majority of which are Tregs, were depleted in the gut mucosa upon acute SIV infection, which correlated with increased gut viral load, inflammation, and intestinal epithelial damage [54]. Long-term ART recovered CD101+CD4+ T cells levels, but these cells expressed higher levels of PD-1, CTLA-4, and TIGIT compared to CD101−CD4+ T cells, thereby potentially promoting HIV latency [54]. On the other hand, HIV infected macrophages have been found to accumulate in duodenal tissue in people with HIV (PWH) [55,56]. Moreover, CD163+CD206+ macrophages were present in the colon submucosa of SIV-infected macaques and these cells were longer-lived compared to CD163+CD206- cells, which may imply these cells as a potential reservoir [57].

Strategies focused on interfering with the HIV reservoir in the GI tract mainly focus on either preventing migration of HIV-infected CD4+ T cells into this site or restoring Th17 cells to counteract their loss. Blocking of α4β7 integrin has been explored as a therapeutic target, as it allows homing of CCR5+ CD4+ T cells into the gut and GALT. However, a clinical trial investigating anti-α4β7 monoclonal antibody therapy in PWH did not show positive effects on viral rebound upon ART interruption [58]. Moreover, in SIV-infected macaques, this therapy increased expression of CD206 and CD163 on mature duodenal macrophages, which correlated with higher viral loads [59].

Since the integrity of intestinal immune barrier is not fully restored despite prolonged ART and residual microbial translocation and inflammation persist in PWH [47,51], strategies to ameliorate HIV-mediated microbial translocation may have major implications for HIV persistence and associated comorbidities. Administration of metformin in PWH under ART reduced inflammation, viral transcription, and immune infiltration of CD4+ T cells in the colon, including Th17 cells [60]. In addition, a recent study reports that CCR5+CD4+ T cells are more abundant in the intestine of conventional mice compared to germ-free mice, possibly indicating that the existing microbiota partially dictates HIV susceptibility and persistence [61]. Consequently, HIV RNA levels were higher in plasma and tissues of conventional mice humanized mice compared with germ-free humanized mice [61]. Modifying the microbiota may therefore be a helpful strategy to diminish gut HIV-associated persistent inflammation, although no direct changes to immune activation were observed after fecal microbiota transplantation in PWH on ART [62]. Still, microbial interventions may provide perspectives in reducing HIV reservoir maintenance in the GI tract by reducing microbial translocation and inflammation.

CENTRAL NERVOUS SYSTEM

Increasing efforts to study HIV persistence within the central nervous system (CNS) have been made during recent years. In humanized mice, HIV-infected T cells and macrophages migrate from the periphery to the brain, potentially establishing an HIV reservoir at this site [63]. Indeed, HIV may infect several cell types within the CNS, including, astrocytes [64], pericytes [65] and myeloid-resident cells, such as microglia and perivascular macrophages [66].

Recent studies found intact proviral DNA in frontal lobe white matter of viremic HIV patients [67▪▪] and HIV patients on ART [67▪▪,68]. Intact proviral DNA has also been found in the basal ganglia and cerebellum of PWH despite ART, indicating presence of HIV reservoir throughout the brain [69]. SIV/HIV-infected microglia in the brain of macaques and ART-treated PWH were shown to be replication-competent [70]. In humans, these microglia were characterized as CD11b+CD68+TMEM119+ cells.

Though “kick and kill” strategies have dominated the HIV cure efforts, the CNS is protected by the blood–brain barrier (BBB), which limits access of systemic drugs to this compartment. Indeed, distinct selective pressures across the CNS compared to other anatomic sites may impact viral evolution and persistence in cellular reservoirs [71]. To address this issue, novel routes of administration which have direct access to the CNS can be considered, such as intranasal drug delivery [72], as well as new systems based on electromagnetic nanoparticles presenting minimal cytotoxic effects [73]. In this sense, nanoformulation of elvitegravir demonstrated in vitro suppression of HIV-1-infected human monocyte-derived macrophages with improved BBB penetration [74]. Of note, strategies to enhance antiretroviral drug delivery to tissues using nanotechnology have recently been reviewed [75] and will not be addressed in this review. Still, many LRAs, including a histone deacetylase inhibitor like Vorinostat, have demonstrated effective CNS penetration and are currently undergoing clinical trials [66]. However, reactivation of HIV in the brain and elimination of cells within the CNS might be complex and could have detrimental consequences [66]. New models using microglia-containing human cerebral organoids [76] or the use of human postmitotic monocyte-derived microglia-like cells [77], may be of great help to address cure strategies in particular subsets of the CNS and may aid screening for safe and effective strategies to target the CNS-HIV reservoir.

OTHER TISSUES RELEVANT TO HIV PERSISTENCE

Evidence of HIV persistence in most tissues across the human body have been reported [78]. Recent literature shows detection of viral DNA and/or RNA at multiple additional tissue sites [79], including hepatocytes and liver-infiltrating CD4+ T cells [80▪], lung bronchoalveolar lavage samples [81], hematopoietic progenitor cells in bone marrow [82,83], kidney [84], urethra [10,31▪▪], testicles [85] and prostate [3]. Further, inflammation at a particular site, such as the gingival tissue [86] or the CNS [15▪▪], may act as the spark that unmasks persistent reservoirs, likely in the form of antigen-specific CD4+TRM that remained in a particular tissue to provide long-term memory. In this sense, application of ingenol mebutate to treat keratosis in ART-suppressed PWH can disrupt HIV latency in the skin tissue microenvironment in vivo[87]. Thus, the persistent depletion of skin CXCR3+CD4+ TRM observed along with skin disease manifestations, despite reconstitution of CD4+ T cells in peripheral blood of PWH receiving late ART [88▪], is likely accompanied by the establishment of a significant reservoir in cutaneous CD4+ TRM.

The difficulty remains to determine which locations and cell types harbor most of the intact virus that may give rise to viral rebound. The heightened resistance of viruses during viral rebound to IFN-I indicates that they either arise from cryptic reservoirs of highly IFN-I resistant viruses or rapidly evolve at the sites of viral recrudescence [89]. In fact, viral rebound can arise from many different tissue compartments [1–3] and intact proviruses located in multiple tissues as well as presence of identical proviral sequences at distant sites suggests circulation of HIV reservoirs under ART [34▪]. Furthermore, recent evidence also indicates that defective proviral DNA may also produce virions despite ART [90], which will have clinical impact. Together, the challenge remains for assessing the weight of these diverse reservoirs on the overall burden of HIV persistence and, also, for determining the best strategy that can selectively reactivate and eliminate these reservoirs.

STRATEGIES FOR REDUCING HIV PERSISTENCE IN TISSUES

Many types of LRA exist and this subject has been reviewed extensively [91]. However, studies addressing the in vivo effect of LRA in tissue are scarce. Challenges to expose these tissue cellular reservoirs ex vivo may not only depend on cell yield but also on the choice of LRA. Although different subsets from peripheral blood may not respond equally to available LRA [92], cell subsets from different anatomical compartments harboring intact proviruses may also respond differently. Beyond blood, studies considering tissue architecture and cell interactions limiting or favoring reservoirs to reactivate upon stimulation have largely been missing. Lack of data also affects other cell types beyond CD4+ T cells, such as macrophages, which may require specific targeting such as TLR4 stimulation to induce reactivation [10,31▪▪].

Engaging innate immunity through TLR activation is a promising strategy to reach reactivation in multiple anatomic reservoirs. Whereas targeting TLR7 has shown potential [93–95], targeting TLR9 recently demonstrated no added benefit to bNAbs alone in the clinics [96]. Moreover, targeting the inflammasome by inhibiting caspase-1 in HIV-infected humanized mice reduced reservoir formation [97]. Additionally, interventions inducing noncanonical nuclear factor κB (NF-κB) signaling by SMAC mimetics may reverse tissue HIV latency in humanized mice and macaques [98], possibly through involvement of galectin-1-glycan interactions [99] and/or synergy with activator protein 1 (AP-1) [100].

Innate immune recognition of mRNA has an intrinsic latency-reversal activity, which could have an impact on HIV persistence [101]. Even when administered intramuscularly, mRNA vaccines against SARS-CoV-2 are capable of eliciting antigen-specific T cell responses in the lung [102]. Therefore, mRNA vaccine efforts may be exploited for reaching HIV reservoirs at tissue sites. Indeed, a recent clinical trial with a nanoparticle-based vaccine (eOD-GT8) against HIV reported establishment of broadly neutralizing antibodies and antigen-specific T cells in vaccinees [103,104], sparking new efforts combining this nanoparticle vaccine with mRNA technology. Lastly, alternative approaches such as genome wide CRISPR-Cas9 screening may identify new targets [105], as shown for aryl hydrocarbon receptor involved in CD4+ T cell viral replication and tissue residency [106]. Other promising approaches to reach the tissue reservoir combine LRA with another therapy, such as a PKC modulator combined with autologous NK cells [107], trispecific antibodies (N6/aCD3-aCD28) [108], IL-15 superagonist N-803 [109], or the targeting of antiapoptotic and autophagy molecules [110].

As a final point, different tissues comprise various states of local metabolic microenvironments, enforcing tissue-resident immune cells to metabolically adapt to this microenvironment. During HIV infection, the metabolic characteristics of the tissue-resident HIV reservoir may be altered, potentially providing a target for their elimination. Indeed, CD4+ T cells of PWH show higher levels of oxidative phosphorylation (OXPHOS) in blood and tonsillar tissue [111,112] and inhibition of OXPHOS by metformin reduced HIV replication in human CD4+ T cells and humanized mice [112]. Although TRM survive in tissue by relying on the availability of exogenous fatty acids to fuel OXPHOS [113], TEM preferentially use glycolysis [114]. As HIV may especially infect TEM expressing high glycolysis levels, partial blocking of glycolysis in vitro resulted in prevention of HIV infection and selective elimination of infected cells [115]. On the other hand, tissue-resident macrophages utilize fatty acids and glucose as a fuel source for OXPHOS for their homeostasis [116]. However, in vitro latent HIV-infected macrophages may shift to glutamate as an energy source and blocking of glutamate resulted in killing of latent infected but not uninfected macrophages [117]. In urethral tissue of PWH, M4 macrophages contained the main HIV reservoir and glycolysis blocking in these cells lead to control of HIV reactivation [31▪▪]. Together, these data indicate that metabolic interventions specifically targeting latent HIV infected cells may contribute to closing in on reducing the HIV reservoir. This becomes especially promising if tissue-specific metabolic traits of the HIV reservoir are considered.

OTHER CONSIDERATIONS

In contrast to men, in whom the HIV reservoir steadily declines with aging, the HIV reservoir in women is more dynamic. Total HIV DNA (including intact and defective genomes) declines more slowly in women than in men, while the inducible HIV RNA+ reservoir, which is highly enriched in replication-competent virus, increases in women after menopause [118▪]. In this sense, a recent HIV cure trial done exclusively in women, the AIDS Clinical Trials Group study A5366, evaluated the impact of tamoxifen to augment vorinostat-induced HIV RNA expression in postmenopausal women. Regardless of the lack of positive results and limitations of the study, this trial established both the feasibility and necessity of investigating novel HIV cure strategies in women with HIV [119▪].

CONCLUSION

Targeting HIV persistence in tissue reservoirs is a complex but essential step towards achieving an HIV cure. This review emphasizes the need to consider the environment and the nature of cells harboring persistent viruses, as well as its early establishment as part of the viral immune response in multiple anatomic compartments of difficult access. Consequently, new models to study tissue reservoirs and identify the mechanisms governing viral persistence will be critical in developing successful therapeutic strategies directed to these compartments. Meanwhile, advances in the area of cancer, including adoptive T cell therapy, gene editing, bispecific antibodies and anti-HIV chimeric antigen receptor (CAR) T cells, reviewed elsewhere [120], hold promise. Overall, we argue that multifaceted and tissue-specific approaches, including early interventions, represent our best hope for limiting HIV persistence and moving the field closer to the ultimate goal of HIV cure.

Acknowledgements

We would like to thank Dr Maria J. Buzón for a thoughtful review of the manuscript. Figures were created with BioRender.com.

Financial support and sponsorship

This work was primarily supported by grants from the Spanish Health Institute Carlos III (PI20/00160), co-funded by ERDF/ESF, “A way to make Europe”/“Investing in your future”, the Fundació La Marató TV3 (202112–30 FMTV3) and the Gilead fellowships GLD21/00049.

Conflicts of interest

There are no conflicts of interest.

REFERENCES AND RECOMMENDED READING

Papers of particular interest, published within the annual period of review, have been highlighted as:

▪ of special interest

▪▪ of outstanding interest

REFERENCES 1. Chaillon A, Gianella S, Dellicour S, et al. HIV persists throughout deep tissues with repopulation from multiple anatomical sources. J Clin Invest 2020; 130:1699–1712. 2. De Scheerder MA, Vrancken B, Dellicour S, et al. HIV rebound is predominantly fueled by genetically identical viral expansions from diverse reservoirs. Cell Host Microbe 2019; 26:347–358. e7. 3. Sun W, Rassadkina Y, Gao C, et al. Persistence of intact HIV-1 proviruses in the brain during antiretroviral therapy. Elife 2023; 12:RP89837. 4▪. Cole B, Lambrechts L, Boyer Z, et al. Extensive characterization of HIV-1 reservoirs reveals links to plasma viremia before and during analytical treatment interruption. Cell Rep 2022; 39:110739. 5▪▪. Gantner P, Buranapraditkun S, Pagliuzza A, et al. HIV rapidly targets a diverse pool of CD4(+) T cells to establish productive and latent infections. Immunity 2023; 56:653–668. e5. 6. Shahid A, MacLennan S, Jones BR, et al. The replication-competent HIV reservoir is a genetically restricted, younger subset of the overall pool of HIV proviruses persisting during therapy, which is highly genetically stable over time. Res Square 2023; doi: 10.21203/rs.3.rs-3259040/v1. 7. Tokarev A, Machmach K, Creegan M, et al. Single-cell profiling of latently SIV-infected CD4(+) T cells directly ex vivo to reveal host factors supporting reservoir persistence. Microbiol Spectrum 2022; 10:e0060422. 8. Abrahams MR, Joseph SB, Garrett N, et al. The replication-competent HIV-1 latent reservoir is primarily established near the time of therapy initiation. Sci Transl Med 2019; 11:eaaw5589. 9. Cantero-Perez J, Grau-Exposito J, Serra-Peinado C, et al. Resident memory T cells are a cellular reservoir for HIV in the cervical mucosa. Nat Commun 2019; 10:4739. 10. Ganor Y, Real F, Sennepin A, et al. HIV-1 reservoirs in urethral macrophages of patients under suppressive antiretroviral therapy. Nat Microbiol 2019; 4:633–644. 11. Packard TA, Schwarzer R, Herzig E, et al. CCL2: a chemokine potentially promoting early seeding of the latent HIV Reservoir. mBio 2022; 13:e0189122. 12▪. Kunzli M, Masopust D. CD4(+) T cell memory. Nat Immunol 2023; 24:903–914. 13▪. Poon MML, Caron DP, Wang Z, et al. Tissue adaptation and clonal segregation of human memory T cells in barrier sites. Nat Immunol 2023; 24:309–319. 14. Genesca M, Skinner PJ, Bost KM, et al. Protective attenuated lentivirus immunization induces SIV-specific T cells in the genital tract of rhesus monkeys. Mucosal Immunol 2008; 1:219–228. 15▪▪. Lisco A, Lange C, Manion M, et al. Immune reconstitution inflammatory syndrome drives emergence of HIV drug resistance from multiple anatomic compartments in a person living with HIV. Nat Med 2023; 29:1364–1369. 16▪. Reeves DB, Bacchus-Souffan C, Fitch M, et al. Estimating the contribution of CD4 T cell subset proliferation and differentiation to HIV persistence. Nat Commun 2023; 14:6145. 17. Bosque A, Famiglietti M, Weyrich AS, et al. Homeostatic proliferation fails to efficiently reactivate HIV-1 latently infected central memory CD4+ T cells. PLoS Pathogens 2011; 7:e1002288. 18. Chomont N, DaFonseca S, Vandergeeten C, et al. Maintenance of CD4+ T-cell memory and HIV persistence: keeping memory, keeping HIV. Curr Opin HIV AIDS 2011; 6:30–36. 19. Simonetti FR, Zhang H, Soroosh GP, et al. Antigen-driven clonal selection shapes the persistence of HIV-1-infected CD4+ T cells in vivo. J Clin Invest 2021; 131:e145254. 20. Vandergeeten C, Fromentin R, DaFonseca S, et al. Interleukin-7 promotes HIV persistence during antiretroviral therapy. Blood 2013; 121:4321–4329. 21▪▪. Dufour C, Richard C, Pardons M, et al. Phenotypic characterization of single CD4+ T cells harboring genetically intact and inducible HIV genomes. Nat Commun 2023; 14:1115. 22▪. Clark IC, Mudvari P, Thaploo S, et al. HIV silencing and cell survival signatures in infected T cell reservoirs. Nature 2023; 614:318–325. 23▪. Sun W, Gao C, Hartana CA, et al. Phenotypic signatures of immune selection in HIV-1 reservoir cells. Nature 2023; 614:309–317. 24. Chomont N, El-Far M, Ancuta P, et al. HIV reservoir size and persistence are driven by T cell survival and homeostatic proliferation. Nat Med 2009; 15:893–900. 25. Perreau M, Savoye AL, De Crignis E, et al. Follicular helper T cells serve as the major CD4 T cell compartment for HIV-1 infection, replication, and production. J Exp Med 2013; 210:143–156. 26▪. Wu VH, Nordin JML, Nguyen S, et al. Profound phenotypic and epigenetic heterogeneity of the HIV-1-infected CD4(+) T cell reservoir. Nat Immunol 2023; 24:359–370. 27. Astorga-Gamaza A, Grau-Exposito J, Burgos J, et al. Identification of HIV-reservoir cells with reduced susceptibility to antibody-dependent immune response. eLife 2022; 11:e78294. 28. Clain JA, Rabezanahary H, Racine G, et al. Early ART reduces viral seeding and innate immunity in liver and lungs of SIV-infected macaques. JCI insight 2023; 8:e167856. 29▪▪. Veenhuis RT, Abreu CM, Costa PAG, et al. Monocyte-derived macrophages contain persistent latent HIV reservoirs. Nat Microbiol 2023; 8:833–844. 30. Ng LG, Liu Z, Kwok I, Ginhoux F. Origin and heterogeneity of tissue myeloid cells: a focus on GMP-derived monocytes and neutrophils. Annu Rev Immunol 2023; 41:375–404. 31▪▪. Real F, Zhu A, Huang B, et al. S100A8-mediated metabolic adaptation controls HIV-1 persistence in macrophages in vivo. Nat Commun 2022; 13:5956. 32. Otte F, Zhang Y, Spagnuolo J, et al. Revealing viral and cellular dynamics of HIV-1 at the single-cell level during early treatment periods. Cell Rep Methods 2023; 3:100485. 33. Baiyegunhi OO, Mann J, Khaba T, et al. CD8 lymphocytes mitigate HIV-1 persistence in lymph node follicular helper T cells during hyperacute-treated infection. Nat Commun 2022; 13:4041. 34▪. Dufour C, Ruiz MJ, Pagliuzza A, et al. Near full-length HIV sequencing in multiple tissues collected postmortem reveals shared clonal expansions across distinct reservoirs during ART. Cell reports 2023; 42:113053. 35. Collins DR, Hitschfel J, Urbach JM, et al. Cytolytic CD8(+) T cells infiltrate germinal centers to limit ongoing HIV replication in spontaneous controller lymph nodes. Sci Immunol 2023; 8:eade5872. 36. Ugur M, Schulz O, Menon MB, et al. Resident CD4+ T cells accumulate in lymphoid organs after prolonged antigen exposure. Nat Commun 2014; 5:4821. 37. Pampusch MS, Abdelaal HM, Cartwright EK, et al. CAR/CXCR5-T cell immunotherapy is safe and potentially efficacious in promoting sustained remission of SIV infection. PLoS Pathogens 2022; 18:e1009831. 38. Rahman SA, Billingsley JM, Sharma AA, et al. Lymph node CXCR5+ NK cells associate with control of chronic SHIV i