Advances in brain targeting anddrug delivery of anti-HIVtherapeutic agentsAbeer M Al-Ghananeem†, Michael Smith, Maria L Coronel & Hieu TranSullivan University, College of Pharmacy, Department of Pharmaceutical Sciences, Louisville,
KY, USA
Introduction: Human immunodeficiency virus (HIV) is a neurotropic virus that
enters the central nervous system (CNS) early in the course of infection.
Although antiretroviral drugs are able to eliminate the majority of the HIV
virus in the bloodstream, however, no specific treatment currently exist for
CNS infections related to HIV. This is mainly attributed to the poor penetrabil-
ity of antiretroviral therapy across the blood--brain barrier (BBB), and the
protective nature of the BBB. Therefore, in order to increase the efficacy of
anti-HIV drugs, novel drug delivery methodologies that can exhibit activity
in the CNS are most needed and warranted.
Areas covered: In this review article, the authors discussed the challenges with
delivering drugs to the brain especially under HIV infection pathophysiology
status. Also, they discussed the approaches currently being investigated to
enhance brain targeting of anti-HIV drugs. A literature search was performed
to cover advances in major approaches used to enhance drug delivery to
the brain.
Expert opinion: If drugs could reach the CNS in sufficient quantity by the
methodologies discussed, mainly through intranasal administration and the
utilization of nanotechnology, this could generate interest in previously aban-
doned therapeutic agents and enable an entirely novel approach to CNS
Human immunodeficiency virus (HIV) appears to be harbored in the brain, as indi-cated by the presence of large quantities of unintegrated viral DNA in the brain ofHIV-infected individuals [1]. A wide variety of nervous system disorders occur inpatients with HIV [2,3]. The most common and devastating central nervous system(CNS) complication associated with HIV infection is a progressive dementia, whichis also referred to as dementia [4,5]. Dementia is a metabolic encephalopathy inducedby HIV infection and fueled by immune activation of macrophages andmicroglia [6].
NeuroAIDS is the neurological complications associated with acquired immuno-deficiency syndrome (AIDS).
The exact mechanism of virus entry into the brain is not clearly elucidated; how-ever the resulting infection leads to a number of CNS disorders [7,8]. Currently, nospecific treatment exists for neuroAIDS, which is mainly attributed to the poor pen-etrability of antiretroviral therapy (ART) across the blood--brain barrier (BBB). Theselective permeability of the BBB is due to the distinct morphology and enzymaticproperties of endothelial cells that enable them to form complex tight junctions
with minimal endocytic activity. This provides a physiologicalbarrier that limits the transport of many blood-borneelements such as macromolecules and circulating leukocytesto the brain [9]. Previous studies report that delivery of ARTto the brain is limited especially due to the physical structureof the BBB, presence of efflux pumps and higher expression ofmetabolizing enzymes, which makes BBB an effective barrieragainst many antiretroviral drugs [10]. Therefore, in order toincrease the efficacy of anti-HIV drugs, novel approaches todeliver antiretroviral drugs to the brain are warranted.Brain drug delivery is a challenge in the treatment of dis-
eases such as neurological disorders. Drugs may be adminis-tered directly into the CNS, or indirectly through oral andintravenous administration. However, transport of drugsfrom systemic circulation into the CNS is restricted by thepresence of the BBB and blood--cerebrospinal fluid barrier(BCSFB), which are featured by tight junctions connectingthe cerebral endothelial and epithelial cells of the choroidplexus, respectively. Most charged, hydrophilic, water-soluble substances, and large therapeutic agents are inhibitedor prevented from entering the brain by the BBB [11,12].Accordingly, to resolve the inhibitory nature of the BBB to
some of the ART drugs, there is a dire need for novel thera-peutics that exhibit antiviral activities in the CNS, attenuateinflammatory pathways and enhance drug delivery intothe brain.This review presented an overview of the brain pathophys-
iology and the current advances in delivery of anti-HIV drugs.
An overview of options in delivering anti-HIV medications tothe brain bypassing the BBB including intranasal delivery andsurgical alternatives were provided. The absorption of anti-HIV medications through BBB was discussed utilizing thenanotechnology, structural manipulation as well as throughaltering BBB passage pathways.
2. Brain pathophysiology and anti-HIVtherapeutic agents
2.1 Pathogenesis of HIV infectionHIV is an obligate intracellular ribonucleic acid (RNA) retro-virus that is highly dependent on host cellular functions tosurvive, replicate and proliferate. This virus is transmittedprimarily through blood and body fluids causing AIDS.The virus interacts with a large number of different cells inthe body and is capable of escaping numerous host immunedefenses against it [13]. Although the exact mechanism of virusentry into the brain is not clearly elucidated, a sequence ofevents occurs to achieve survival. HIV surface gp120 glyco-protein will attach to the CD4 receptor located on the hostcell membrane and the interaction of the gp120 proteinand CD4 complex leads to virus--cell membrane fusion medi-ated by transmembrane gp41 protein [14]. Effective survival ofretroviruses depends on exploiting host cell processes withinthe cell. Viral integrase is responsible for integration of areverse-transcribed viral cDNA into the chromosome of theinfected cell which is a crucial step in the early phases ofthe retroviral life cycle. Retroviral integration causes hostcell chromosome damage. Activation of DNA repair canresult in arrest of cell cycle, which provides time for DNArepair. However, if DNA repair system was left unrestored,apoptosis or cell death can occur [15]. Retroviruses differ intheir preferences for sites for viral DNA integration in thechromosomes of infected cells. HIV integrates preferentiallywithin active transcription units [16]. Proper integration byblocking either integrase catalysis or the function of cellularintegration may potentially offset chromatin alteration pre-venting cell apoptosis.
HIV contains a viral envelope surrounded by projectionsformed from the proteins gp120 and gp41. During an activeinfection, the HIV envelope gp120 glycoprotein is shed bythe virus from HIV-infected immune cells [17]. In the CNS,this generates inflammation and oxidative stress that contrib-utes to the development of the AIDS Dementia Complex(ADC). The correlation of the excitotoxic effects ofgp120 to astroglial cells has been demonstrated through lipidperoxidation and altered glutamine release to brain cell dam-age. Maintaining an active balance activity of glutamine syn-thase (GS), the enzyme that metabolizes glutamate intoglutamine in astroglial cells plays a neuroprotective role [18].
There are further suggestions that neuronal injury observedin the brain of AIDS patients is related to excessive influx ofCa2+ associated with the neurotoxic injurious effect inducedby gp120. This mechanism of toxicity involves the activation
Article highlights
. Dementia is the most significant and devastating centralnervous system (CNS) complication associated withhuman immunodeficiency virus (HIV) infection.
. Currently, no specific treatment exists for CNS infectionsrelated to HIV. This is mainly attributed to the poorpenetrability of antiretroviral therapy (ART) across theblood--brain barrier (BBB), and the protective nature ofthe BBB making the brain tissue HIV sanctuary site.
. Various strategies were implemented for enhanced BBBpenetration, including nanotechnology, structuralmanipulation as well as the use of transport/carrier systems. Other strategies for drug delivery to thebrain involve bypassing the BBB such as intranasaladministration and direct invasive methods.
. Research efforts should focus on enhancing the portionof the nasal dose that reaches the CNS.
. Novel developments emerging in the field of polymerscience and nanotechnology provide an option by whichthe obstacles of limited brain entry can be surmounted.
. A better appreciation of the transporters present at thebrain barriers could prove a valuable milestone inunderstanding the limited brain penetration of anti-HIVdrugs in HIV. This also could aid the development ofnew anti-HIV drugs, new nanotechnology templates anddrug structural manipulations, with enhanced efficacy inthe CNS.
This box summarizes key points contained in the article.
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of voltage-dependent Ca2+ channels and N-methyl-D-aspartate(NMDA) receptor-operated channels. These suggestionsoffer avenues for future pharmacological intervention withfocus on clinically tolerated calcium channel antagonists andNMDA antagonists as potential trials in humans with AIDSdementia in the near future [19,20]. Further pathogenesis inHIV-infected patients is explained in Table 1 [21-27].
2.2 Dementia and AIDS in HIVThe term dementia is used to describe a set of symptomsresulting from damages and disorders affecting the brain.The associated symptomatology can be due to the specificregions affected by a multitude of diseases known as neurode-generative diseases. The symptoms presented vary fromdecreasing cognitive functions to behavioral, affective, motorand psychiatric disorders. The terms ADC, and HIV-1 associ-ated dementia (HAD), are used to describe these neurologicaland psychiatric symptoms caused by human immunodefi-ciency virus type 1 (HIV-1) infection [22,25]. HIV-1 is themost common cause of dementia in young adults in theUSA [25,26].
2.3 Brain pathophysiological status and its impact
on anti-HIV drugsThe HIV virus commonly affects the basal ganglia, deep whitematter and cerebral cortex in HIV/AIDS dementia. Patholog-ical features include pervasive reactive astrocytosis, myelin pal-lor, activated resident microglia, infiltration by circulatingmonocytic cells, perivascular inflammation, microglial nod-ules, multinucleated giant cells, dendritic simplification andcell death with both astrocytic and neuronal damage [27].
Despite the current therapeutic management with activeART, neuronal cell death persists and has become a constantproblem affecting the CNS of HIV-infected patients.Although anti-HIV therapy has successfully prevented manyof the former end-stage complications of AIDS leading to
increased survival times, the HAD is still prevalent in treatedpatients as well as attenuated forms of CNS opportunisticdisorders. This suggests that there may be other indirectmechanisms of neuronal injury and loss/death occur inHIV/AIDS as a basis for the dementia which can still beexplored [13].
The anatomical--physiological protective mechanism of theBBB and the BCSFB has garnered interest due to the chal-lenges imposed in therapeutics and adverse outcomes ofdrug interactions. Faced with challenges in altered efficacyand enhanced neurotoxicity of drugs, clinicians are posedwith problems in drug delivery and treatment of diseaseslike AIDS dementia [28].
Recent and past clinical therapeutic trials for the treatmentof dementia have focused on some drugs as adjunct therapy tocurrent anti-HIV therapy. Several recent, large-scale trialsconducted through the AIDS Clinical Trials Group(ACTG) focus on three drugs: minocycline, memantine andselegiline as adjunct therapy to anti-HIV drugs (Table 2).Interestingly, data suggested neuroprotective properties ofminocycline that makes it an excellent candidate for limitingHIV replication within the brain, which has remained a diffi-cult anatomical compartment for therapeutic interventionsfor HIV due to the lack of BBB permeability of many suchdrugs [27].
The BBB and BCFSB are not merely passive barriers butalso play a dynamic role in the expression of influx and effluxtransporters and drug-metabolizing enzymes. Studies indicatethat efflux transporters such as P-glycoprotein (P-gp) play animportant role in drug delivery [29]. Furthermore, chemicalinhibition of transporters predicts lesser magnitude ofexpected drug--drug interaction in human BBB but may beclinically significant [29]. Defining the impact of such drug--drug interactions is important for improving efficacy of drugsused in the treatment of CNS diseases while minimizingneurotoxicity.
Table 1. Pathogenesis and neurodegeneration mechanisms in HIV infection.
Pathogenesis in HIV Levels in HIV infection Mechanism of neurodegeneration
Direct effect of HIV infection by proteins Increase HIV-1 gp120 Stimulate nitric oxide synthaseIncrease release of arachidonic acidInhibition of glutamate uptake by astrocytes and neurons
Increase HIV-1 Tat Disruption of the BBBInfiltration of inflammatory cells into the CNSIncreases cytokines and chemokines
HIV-1 Vpr Induce apoptosis of human neuronal-precursor cellsActivation of caspase-8
Indirect consequence of infectionthrough inflammation
Increase chemokineand cytokine
Leukoencephalopathy and neuronal apoptosisGlutamate-associated neurotoxicityTNF-a-associated damage to the BBB increasingentry of HIV proteins and cytokines
BBB: Blood--brain barrier; CNS: Central nervous system; gp120: Glycoprotein 120; HIV: Human immunodeficiency; HIV-1: Human immunodeficiency virus type 1;
Tat: Transcriptional transactivator; Vpr: Viral protein R.
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3. Delivering anti-HIV medication to brainbypassing the BBB
3.1 Intranasal drug deliveryDelivery of drugs into the brain via intranasal administrationhas gained increased attention in recent years. Intranasal deliv-ery is a non-invasive method that targets drugs to the brainand spinal cord along olfactory and trigeminal neural path-ways, bypassing the BBB and minimizing systemic exposureand side effects. Nasal dosage forms can be self-administeredby the patient and do not need a healthcare provider like par-enteral administration. Furthermore, the nasally applied drugis immediately absorbed through the nasal mucosal mem-branes, largely avoiding hepatic first-pass metabolism. More-over, a part of the absorbed drug is distributed directly tothe CNS as well as via blood flow.In addition to the systemic absorption of therapeutic agents
after intranasal administration, direct delivery of a wide varietyof therapeutic agents to the CNS following intranasal adminis-tration, as well as the therapeutic benefit of intranasal drug deliv-ery has been demonstrated by the authors’ group andothers [30-36]. The direct anatomical connection exists betweenthe nasal cavity and the CNS makes it possible to deliver sometherapeutic agents into the CNS by circumventing the BBB,which provides the basis for the development of therapeuticagents for intranasal administration. Potential mechanisms fornose-to-brain drug transport are thought to involve the trigemi-nal nerve receptors and/or one or a combination of two generalmechanisms namely: axonal transport through the olfactorynerve and absorption across the olfactory sustentacular epithelialcells [36]. Thus, the poor penetration of antiviral agents into theCNS may potentially be overcome by intranasal administration.The nasally administered gp160-HVJ-liposome to normal
mice has shown to be a good immunization tool that induces
necessary Ag-specific immune responses at different stages ofHIV [37]. Furthermore, brain tissue, CSF and olfactory bulbwere shown to have higher concentrations of intranasal didan-osine (2¢,3¢-dideoxyinosine, ddI) following nasal administra-tion when compared with intravenous administration. Theresults suggest that ddI can reach CNS compartments via adirect pathway initiating in the nasal cavity that does notinclude systemic circulation [38].
Intranasal administration of zidovudine (3¢-azido-3¢-deoxy-thymidine, AZT) in rats showed rapid absorption and highCSF concentrations of AZT. Furthermore, nasal delivery ofAZT co-administered with probenecid resulted in high CSFconcentration of AZT [39]. Accordingly, the nasal route wassuggested as a non-oral and non-parenteral dosage form choicefor AIDS treatment to patients with CNS dysfunctions.
The CSF and brain levels of AZT after intranasal adminis-tration of a thermo-reversible gel formulation were greaterthan those attained after intravenous injection. The pharma-cokinetic and brain distribution levels revealed that a polarantiviral compound such as AZT could preferentially transferinto the CSF and brain tissue via an alternative pathway,possibly olfactory route after intranasal administration [40].
Depending on the therapeutic agent physicochemical prop-erties, sometimes nasal administration of therapeutic agentsdoes not result in high levels in CNS. The extent of brain dis-tribution of stavudine after intranasal administration in ratswas comparable or sometimes less than that following sys-temic intravenous administration [41]. It seems that the directtransport pathway from the nasal cavity to brain did not con-tribute significantly to the overall brain exposure of stavudine.
In summary, although intranasal administration has beenused for brain delivery of some therapeutic agents, the effi-ciency of this route in human is still uncertain due to the smallolfactory mucosa area with respect to rodents or dogs. Note
Table 2. Potential effects of adjunct therapy on HIV pathology per studies reported from the ACTG [49].
Yes Transdermal administrationof selegiline may avoidtoxicities
Disappointing results in arecent short-term clinicaltrial on transdermal
ACTG: AIDS Clinical Trials Group; BBB: Blood--brain barrier; HIV: Human immunodeficiency; MAOIB: Monoamine oxidase type B inhibitor;
NMDAR: N-methyl-D-aspartate.
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that the olfactory area constitutes 50% of the nasal cavity inrat, but approximately 3% in human [36]. Thus, the transportmechanism to CNS through this pathway requires furtherclinical investigation to better understand the criteria definingtherapeutic agents’ properties needed for better absorptionand direct delivery to brain.
3.1.1 Surgical optionsInvasive surgical approaches for drug delivery are a means tobypass the BBB that carries risks for the patients withoutassurance of adequate drug distribution. This entails specialdelivery methods that involve direct introduction of the ther-apeutic substances into the brain or into the CSF such as neu-rosurgical procedures, implantation of cells or tissues or theuse of special devices [42-44].
Manipulation of catheters into the CSF compartment (lumbarsubarachnoid space, cisterns and ventricles) during angiographyallows direct injection of contrast media or therapeutic substan-ces into the CNS [45]. This enables a precise placement of electro-des or probes in the brain with minimal invasion achievingsuccessful drug distribution. Direct convection delivery can alsobe used to deliver an infusate and distribute macromolecules ina predictable, homogeneous manner over significant volumes [46].Interstitial delivery of drugs to the brain from biological tissues isa promising strategy to achieve the desired therapeutic agentthrough the implantation of genetically engineered cells thatsecretes the required drug [47-49]. Chimeric peptide technology,wherein a non-transportable drug is conjugated to a BBB trans-port vector holds as much promise for future success [50,51].Intraventricular and intrathecal route uses an Ommaya reser-voir implanted subcutaneously in the scalp and connected tothe ventricles of the brain. Anti-HIV drugs are introduced sub-cutaneously into the implanted reservoir, which subsequentlydelivers the administered drugs to the cerebral ventricles, bymanual compression of the reservoir through the scalp.A smaller dose may be used to offset any potential toxicity.Unfortunately, this delivery method has not been favored dueto its slow rate of drug distribution with the CSF and theincrease in intracranial pressure. Also, it carries high risks forhemorrhage, CSF leaks, infections and neurotoxicity [52-54].
4. Absorption of anti-HIV therapeutic agentsthrough BBB
4.1 Formulation approaches through nanotechnologyDue to their small size range, nanoparticle systems have beenproposed as potential brain targeting drug delivery sys-tems [55,56]. Recent advances in nanotechnology have providedpromising solutions to brain targeting challenge. Orallyadministered stavudine-loaded solid lipid nanoparticles wereeffectively targeted to cells of the reticuloendothelial system(RES) and brain. The solid lipid nanoparticles of stavudineshowed an 11-fold increase in stavudine brain uptake in com-parison with orally administered stavudine solution [57]. Fur-thermore, solid lipid nanoparticles were reported to enhance
the delivery of atazanavir by human brain endothelial cellline [58]. Nanoparticle drug delivery systems have beenincreasingly used to improve the efficiency of antiretroviraldrugs such as saquinavir [59-62]. Recent studies show that thespecificity and efficiency of antiretrovirals delivery can be fur-ther enhanced by using nanotechnology with specific braintargeting, cell penetrating ligands or ATP-binding cassette(ABC)-transporters inhibitors [63].
Several nanotechnology systems ranging from the moreestablished systems, for example, polymeric nanoparticles,solid lipid nanoparticles, liposomes, micelles to the newer sys-tems, for example, dendrimers, nanogels, nanoemulsions andnanosuspensions have been studied for the delivery of CNStherapeutics. Many of these nanomedicines can be effectivelytransported across various in vitro and in vivo BBB modelsby endocytosis and/or transcytosis, and demonstrated earlypreclinical success for the management of CNS conditionssuch as HIV encephalopathy [38,63-65].
Significant enhancement in ddI brain tissue concentrationwas observed when administering the drug nasally inchitosan-loaded nanoparticles in comparison with nasallyadministered ddI solution [38].
These nanotechnology studies suggest considerable oppor-tunities to enhance delivery of therapeutic agents to theCNS for HIV therapy.
4.2 Targeted delivery through structural
manipulationsIn order to overcome the bioavailability issue associated withantiretroviral agents in the CNS, a number of structural mod-ifications to nucleoside reverse transcriptase inhibitors (NRTIs)and protease inhibitors (PIs) have been created in order toimprove BBB penetration. The antiviral AZT is classified as aNRTI which was the first drug approved for the treatment ofAIDS. On drug administration, monitored levels of AZT inthe brain are low relative to the periphery [66]. This phenome-non is explained through a number of mechanisms; AZT hasbeen shown to cross the BBB by a carrier-mediated mechanismand a passive diffusion process [67,68]. Additionally, AZT is sus-ceptible to active efflux from the brain into the blood throughP-gp-mediated efflux [69-71]. AZT has undergone a number ofmodifications with the intent of increasing CNS penetration(Table 3). A high volume of promoieties have been introducedto 5¢-hydroxyl group connected by a labile ester linkage [72-78].Some notable promoieties introduced are retinoic acid whichalone has the capacity to inhibit HIV replication [73]. Unfortu-nately, after subsequent prodrug conversion an additive effectwith AZT was not observed but it contained the highestH9 cellular uptake in that series of prodrugs. In the same study,amino acid ester prodrugs were synthesized with purpose ofenhanced penetration into the CNS possibly through theL-amino acid transporter, the isoleucine prodrug displayedthe highest lipid partition coefficient and not surprisinglydisplayed higher H9 cell uptake relative to AZT. A glycosylphosphotriester prodrug showed improved brain delivery of
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AZT, and is metabolized to a monophosphate metabolitebypassing the need for viral kinase activation [74]. Modificationsto the 5,6-olefin bond of AZT yielding a series of 5-halo-6-alk-oxy (or azido) derivatives which led to enhanced lipophilicityand a comparably higher distribution in the brain relative tothe parent compound [79,80]. Unfortunately, increasing lipophi-licity is not the only hurdle to overcome, there are a number ofmetabolic enzymes located at brain endothelial cells which havebeen referred to as an enzymatic or ‘metabolic’ BBB [81]. Aden-osine deaminase (ADA) and purine nucleoside phosphorylase(PNP) are two significant enzymes located in this region andare involved in purine metabolism. One notable prodrug deriv-ative to ddI, 2¢-b-fluoro-2¢,3¢-dideoxyadenosine (F-ddA) is anacid stable analog of 2¢,3¢-didesoxyadenosine (ddA), which isdeaminated by ADA to the active 2¢-b-fluoro-2¢,3¢-dideoxyino-sine (F-ddI) [82,83]. In addition to improve lipophilicity,F-ddA exploits the enzymatic BBB resulting in an eightfoldhigher brain/plasma ratio compared with equivalent F-ddIinfusion [84].In general, prodrug strategies involve the introduction of a
lipophilic moiety connected by a labile linkage in order toenhance diffusion across biological membranes. Chemicalmodifications of antiretroviral agents with the intent ofenhanced uptake via protein transporters at the BBB areanother viable strategy [85-96].A class of guanidine-rich molecular transporters has been
synthesized that possess a high molecular weight, good watersolubility and can penetrate the mouse brain in vivo [85].More specifically, an eight guanidine (G8) molecular trans-porter was shown to be the most effective at penetrating theBBB. Consequently, a covalently linked AZT with sorbitolG8 (Table 3) was synthesized [86]. Tissue distribution studiesin mice indicated the modified AZT possesses an affinitytoward both the brain and liver.Platelet-activating factor (PAF) has been identified as an
HIV-induced neurotoxin [87,88]. One possible route of neuro-toxicity is through a PAF-activated increase in Ca2+ leading toenhanced excitatory neurotransmission through glutamaterelease [89]. These findings prompted the investigation ofPAF antagonists for potential antiviral activity resulting in atrisubstituted piperazine, PMS-601 possessing both anti-HIV-1 effects as well PAF antagonist activities in the micro-molar range in vitro [90]. Additionally, it was shown to havethe capacity to cross the BBB in rat models using the in situbrain perfusion method, further SAR studies were completedresulting in analogs with enhanced dual activities [91,92]. Thesestudies display the potential to both penetrate the viral poolsin the brain as well as attenuate neurotoxicity.Attempts have also been made to utilize the body’s own
nutrient carrier transport through conjugation of antiviralswith amino acids or D-glucose [93-95]. Amino acid conjugatessuch as L-valyl, L-phenylalanyl and L-leucyl esters of indinavirand saquinavir showed an enhanced transport across theCaco-2 cell monolayer, however D-glucose conjugates showeda decrease in absorption [96]. Further investigations of the
valine derivative of indinavir showed no brain distributionand decreased oral bioavailability relative to indinavir [97].
4.3 Altering BBB passage pathwaysABC transporters such as P-gp and multidrug resistance-associated proteins (MRP) are efflux membrane transporterswith a broad capacity of xenobiotic substrates. They are foundin various tissues of the body including the gastrointestinal tract,liver and kidney [98]. P-gp is expressed at high levels in the braincapillaries and is the primary cause for efflux of antiretrovirals outof the CNS [99,100]. Thus, inhibition of P-gp has advantageousclinical implications in the treatment of NeuroAIDS. Com-monly, PIs (a P-gp substrate) are co-administered with a lowdose of ritonavir which functions to block both P-gp as well ascytochrome P450 metabolizing enzymes [101]. In fact, ritonavirdisplays a higher potency of inhibition of P-gp relative tomultidrug resistance reversing agent SDZ PSC833 [102].
Probenecid is a uricosuric agent and a known inhibitor ofthe organic anion acid transporter in the renal tubule. Notonly does it act in the kidney but also has been demonstratedto block transport of organic acids from the CNS to plasmasuch as penicillin, pantothenic acid and leukotrieneA4 [103-105]. Probenecid administered in combination withAZT shows an enhanced distribution in the brain, presum-ably through a reduced rate of drug efflux [106,107]. Combina-tions of ddI and probenecid also displayed an enhancement inCNS uptake [108]. Looking at the efflux rate of both AZT andddI after microinjection into the brain, demonstrated AZTefflux from the brain was inhibited by ddI and probenecid.Additionally, the efflux rate of ddI was inhibited by AZTand probenecid [71]. This provides supporting evidence thatAZT and ddI are transported from the brain to blood acrossthe BBB via a probenecid-sensitive efflux transporter. Amphi-philic block copolymer (Pluronic P85) has been under investi-gation for the inhibition of efflux transporters. P85 consists oftwo chains of hydrophilic polyethylene oxide groups surround-ing a hydrophobic polypropylene oxide moiety. Initially beingused in cancer resistance studies, P85 was also shown to inhibitP-gp-mediated drug efflux in bovine brain microvessel endo-thelial cells providing a possible route to improve CNS drugpenetration [109]. When used in combination with antiretroviraldrugs, P85 increased the CNS biodistribution of the AZT/lam-ivudine/nelfinavir cocktail in a mouse model of HIV encepha-litis (HIVE) [110]. The immunodeficient mice were inoculatedwith HIV-infected human monocyte-derived macrophages(MDM) into the basal ganglia and then treated with P85,ART or P85 and ART. A significant decrease in the HIV-infected MDM in all treatment groups relative to the controlwas observed, it is important to note that P85 alone containedantiretroviral effects [110]. Thus, P85 was hypothesized to havetwo mechanisms of P-gp inhibition: decreasing ATP levels inbovine brain endothelial cell monolayers and can compromisethe lipid bilayer causing a destabilization of P-gp. This copoly-mer also effectively inhibits the interaction between saquinavirand nelfinavir in cellular accumulation studies, Pluronic
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Table 3. A summary of anti-HIV drugs chemical modifications to improve their CNS bioavailability.
Parent drug Derivatives Therapeutic effect
AZT
O
N3
RO
N
NH
O
O5′-OH Promoieties
Retinoic acidCH3 CH3 CH3
CO2R
CH3CH3
Highest H9 cell uptake in thisseries, however a sixfold increasein cytotoxicity observed relative toAZT. Retinoic acid has been shownto have antiviral activity [66]
Isoleucine
H3C
NH2
OR
OCH3
Isoleucine displayed the highestH9 cell uptake relative to otheramino acid promoieties inthis series [66]
G8 molecular transporter
OR2
R2O
OR2
OR2
O
O
NH25
O
R2 =
O
N5
NH
NH2
HN
HN NH2
NH
O
OR
O
Covalent linkage of a sorbitol-G8transporter displayed enhancedwater solubility and improvedAZT BBB penetration in mousebrain [78]
Glycosylphosphotriester
P ORO
O
O
O
O
HO
OH
OHOHOHOH
Displayed high concentrations ofAZT in vitro via glucosyl dicholphosphate transport, enhancedwater solubility, bypasses viralkinase activation [67]
Modified olefin of AZT
O
N3
HO
N
NH
O
O
RA
RB
RA = Br, Cl, IRB = OMe, OEt, O(CH2)xCH3, N3
Addition to the olefin yielding aclass of 5-halo-6-alkoxy (or azido)AZT lead to enhanced logPand CNS biodistribution,5-bromo/5-iodo analogs mayserve as prodrugs due toregeneration of the olefin [73]
ddI
N
NHN
N
O
OHO
F-ddA
N
O
HN
OH2N
HN O
OR
O
NH
Addition of the 2¢-b-fluoroenhances acid stability, while6-amination yields a prodrug thatundergoes conversion by ADA toF-ddI leading to an eightfoldhigher brain/plasma ratio relativeto F-ddI infusion [75-77]
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F88 and F127 (more hydrophilic copolymers) were also testedbut failed to be active [111].Recently, dimers of abacavir (Table 3) were engineered with a
disulfide linker that was shown to be a potent P-gp inhibi-tor [112]. The prodrug serves a unique dual role of both P-gpinhibition and antiretroviral properties on metabolism. Underthe reducing conditions of the cytosol, the disulfide linker iscleaved ultimately releasing the active monomer. The derivativeAbaS2Me4 displayed the highest activity and a 700-foldincrease in potency over abacavir. Introduction of methyls adja-cent to the carbonyl most likely leads to a reduced metabolismby plasma esterase contributing to the enhanced potency. Thismethod paves the way for varied ART combination in a similardimer which can lead to enhanced CNS penetration, decreaseddosing requirements and a possible lowering of the pill burdenof patients fighting HIV. Targeting P-gp is a viable strategythat shows promise in increasing the brain concentration ofantiretroviral mediations. In fact, the initial thought of in-creasing the drug lipophilicity to enhance drug concentrationsin the brain is possibly counterintuitive. P-gp has beendescribed as a ‘hydrophobic vacuum cleaner’ resulting inincreased removal of lipophilic drugs from the CNS, thusP-gp inhibition combination has clinical significance [112].
5. Conclusion
The restricted entry of anti-HIV drugs to the CNS due to thepresence of BBB hinders the penetration of anti-HIV drugs
into the brain, promoting viral replication, the developmentof drug resistance and ultimately sub-therapeutic concen-trations of drugs reaching the brain, leading to therapeuticfailure. A better appreciation of the transporters present atthe brain barriers could prove a valuable milestone in under-standing the limited brain penetration of anti-HIV drugs inHIV. This also could aid the development of new anti-HIV drugs, new nanotechnology templates and drug structuralmanipulations, with enhanced efficacy in the CNS.
Furthermore, a better understanding of methods thatdeliver anti-HIV drugs to CNS bypassing the BBB is also war-ranted for better control of neurological complications associ-ated with HIV.
6. Expert opinion
The treatment of HIV/AIDS dementia continues to be chal-lenging as we pursue ways to achieve the delivery of optimallevels of drug therapy in the brain for the cure of this debilitat-ing disease. Of further importance is the possibility that theCNS may serve as a sanctuary for the virus after its systemiceradication. The major challenge to CNS drug delivery is theBBB, which limits the access of drugs to the brain substance.Currently approved drugs for the treatment of HIV infectionsmay not attain sufficient drug concentrations in the CNS, andthus may not completely suppress viral replication in the brainover a sustained period of time.
During the last years, considerable efforts were focused onthe field of brain-targeted drug delivery. The ultimate goals
Table 3. A summary of anti-HIV drugs chemical modifications to improve their CNS bioavailability (continued).
Parent drug Derivatives Therapeutic effect
Saquinavir
N
N
N
N
HN
H2N
OH
R = ester/carbamate linked glucoseR = ValR = PheR = Leu
These amino acid prodrugsdisplayed enhanced transportacross Caco-2 monolayer. Furtherstudies of the valine analogdisplayed no brain distributionin vivo. D-Glucose analogsdisplayed a decrease in absorption,additionally ester linkedpromoieties displayed enhancedanti-HIV activity relative to thecarbamate [86-90]
Abacavir
N
NN
N
H2N
OF
HO
Abacavir dimer
N
N
N
N
HN
NH2
O
O
S
N
N
N
N
HN
H2N
O
O
S
Dimeric prodrugs of abacavirdisplay potential for P-gp inhibitionat the BBB and revert to the activemonomer. This presents a platformthat could have application intherapies involved with limitationsof P-gp drug efflux [105]
of those research efforts were mainly to deliver the anti-HIVtherapeutic agents in therapeutically effective concentrationsto the brain. Various strategies were implemented forenhanced BBB penetration, including nanotechnology, struc-tural manipulation as well as the use of transport/carrier sys-tems. Other strategies for drug delivery to the brain involvebypassing the BBB such as intranasal administration anddirect invasive methods. Both strategies have advantages anddisadvantages (Table 4). Although, each strategy may offermany intriguing possibilities for brain delivery and targeting,but only some have reached the phase where they can providesafe and effective human applications.
It is important to consider not only the net delivery of theagent to the CNS, but also the ability of the agent to accessthe relevant target site within the CNS. Site-target and theuse of targeting enhancement factors can be used to quantita-tively assess the effectiveness from a pharmacokinetic perspec-tive of chemical delivery systems. However, from drugdevelopment and regulatory aspects, pharmaceutical compa-nies usually tend toward designing improved formulationsof well-known therapeutic agents that are already in the mar-ket, rather than synthesizing a new chemical entity. Note thatany slight structural manipulation, even simple ester-pro-drugs, will make the therapeutic agent considered as a newchemical entity. The latter would require an extensive regula-tory efforts through new drug application (NDA) necessitat-ing a full safety and efficacy assessment from preclinical andclinical prospective.
A fruitful area of future research will be to determine ifthere are any systems that can target drug delivery to the brain.Novel developments emerging in the field of polymer scienceand nanotechnology provide an option by which the obstaclesof limited brain entry can be surmounted. Thus, the use ofnanotechnology based on Food and Drug Administration(FDA)-approved polymeric carriers would be an attractive
approach to deliver known anti-HIV drugs to the CNS. Theplethora of transporters expressed at the brain barriers all actas selective gatekeepers, and this remains a major obstacle forantiviral therapy. Future directions in nanotechnology as atool to enhance anti-HIV drugs bioavailability in CNS shouldconsider nanocarriers with ligands specific for certain BBBtransports to overcome this obstacle. In the authors’ opinion,this is a milestone enhancement in delivering therapeuticagents including anti-HIV drugs to CNS. Future directionshould focus on two main directions in utilizing nanotechnol-ogy for brain targeting: i) allowing drug trafficking by endocy-tosis (non-specific or receptor-mediated) and ii) blockingdrug efflux transporters. Thus, a better appreciation of BBBtransporters is warranted for successful drug delivery.
Furthermore, applying anti-HIV therapeutic agents encap-sulated into nanoparticles could potentially improve the directCNS delivery via the nasal cavity. Intranasal drug deliveryholds a promising future in brain targeting. However, a chal-lenge of the intranasal administration to target the brain, thatneeds to be in mind during drug development effort, is thatthe quantities of drug administered nasally that have beenshown to be transported directly from nose-to-brain are rela-tively low. Research efforts should focus in enhancing the por-tion of the nasal dose that reaches the CNS. If drugs couldreach the CNS in sufficient quantity by this route, it couldgenerate interest in previously abandoned drug compoundsand enable an entirely novel approach to CNS drug delivery.
Currently, there is no approved nanotechnology-based CNSdrug available; the future for such nanotechnology-baseddelivery system developments is promising. A thorough under-standing of BBB transporter, its location, level of expressionand transport potency is needed to help improve the treatmentof HIV infection in the brain.
Interestingly, there are many nanotechnology anti-HIVplatforms tested but most are still in preclinical research
Table 4. Main advantages and disadvantages of the two drug delivery strategies (absorption through BBB or
bypassing the BBB) used for anti-HIV drug delivery to CNS.
Method Advantage Disadvantage
Bypassing the BBB Bypass hepatic first-pass metabolismReduction of systemic side effectsRapid absorption and onset of action
Devices needed for drug delivery
Surgical options Targeted drug delivery andCNS abundanceControlled and sustained drug delivery
InvasiveSurgical needsHigh risk small site for absorption
Intranasal drug delivery Ease of administrationNon-invasive
Mucociliary clearancePotential nasal irritationCNS drug bioavailability depends on drugabsorption from nasal cavityDose size limitation
Absorption through BBB Ease of administrationPotential technology enhancement withspecific receptors ligands
Systemic exposureLow CNS bioavailabilityOnset of action may be delayedPotential side effects
BBB: Blood--brain barrier; CNS: Central nervous system; HIV: Human immunodeficiency.
Advances in brain targeting and drug delivery of anti-HIV therapeutic agents
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with one immunotherapy-based product making it to clinicaltrials. However, the authors expect to see in the futurenanotechnology-based anti-HIV marketed products, as theFDA expressed interest in such technology translated in open-ing multi-nanotechnology research centers. Research done inthis field holds up hope, despite the BBB shielding challenges.Future research should focus on achieving brain delivery ofanti-HIV therapeutic agents in a safe, efficient and yet cost-effective manner.
Acknowledgment
The authors would like to thank Amber Jeffries for heradministrative assistance in this review article.
Declaration of interest
The authors state no conflict of interest and have received nopayment in preparation of this manuscript.
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AffiliationAbeer M Al-Ghananeem† PhD, Michael Smith,
Maria L Coronel & Hieu Tran†Author for correspondence
