Development of Non-Nucleoside Reverse Transcriptase Inhibitors for Anti-HIV Therapy

7/31/2019 Development of Non-Nucleoside Reverse Transcriptase Inhibitors for Anti-HIV Therapy

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Anti-Infective Agents in Medicinal Chemistry, 2008, 7, 101-117 10

1871-5214/08 $55.00+.00 2008 Bentham Science Publishers Ltd.

Development of Non-Nucleoside Reverse Transcriptase Inhibitors for Anti-HIV Therapy

Christer Sahlberg* and Xiao-Xiong Zhou

Medivir AB, Lunastigen 7, S-141 44 Huddinge, Sweden

Abstract: TheNNRTIs play an important role in the present therapy against HIV/AIDS. This review discusses the basicprinciples in the development of NNRTIs for HIV therapy. It also summarizes the NNRTIs in clinical use and the majorseries of NNRTIs in development phases. The authors intend to provide an overview of the NNRTI research and to eluci-date some important factors in directing the future in the field such as genetic barrier, QD dosing, safety profile andcombination with other anti-HIV agents. Despite the enormous progress that has been achieved in the NNRTI field in thepast two decades, the present clinical pipeline appears to be insufficient to tackle the huge medical need. The efforts offinding new NNRTIs are certainly much motivated and can be highly rewarding.

INTRODUCTION

AIDS, caused by the HIV virus, is one of the worlds

leading causes of death with a major medical and economicimpact on society. The current combination therapy of threeor more drugs is increasing the survival of HIV-infected pa-tients by many years and provides an improved quality oflife. The present therapies are mainly based on the inhibitionof two key viral enzymes: HIV reverse transcriptase (RT)and HIV protease (PI) as well as the inhibition of viral fu-sion. Based on the chemical structures and the inhibitorymechanism, the RT inhibitors can be divided into nucleosideand nucleotide RT inhibitors (NRTIs) and non-nucleosideRT inhibitors (NNRTIs) which bind to an allosteric site ofHIV-1 RT located about 10 away from the catalytic site.

The NNRTIs play an important role in current anti-HIVtherapy as a part of a successful combination therapy. Dif-

ferent aspects of NNRTIs have recently been reviewed suchas; NNRTIs in general [1-7], specific NNRTIs [8-13], resis-tance issues [14,15], x-ray and binding of NNRTIs [16,17],clinical use of NNRTIs [18-21] and toxicity issues withNNRTIs [22-25]. The present review intends to provide anoverview of the landscape of NNRTI research, to highlightsome important principles and aspects in the development ofNRTIs. The review is divided into three parts (i) HIV-1 RTand its inhibition (ii) NNRTIs in clinical use and (iii)NNRTIs in development.

RT AND ITS INHIBITION

HIV-1 RT

HIV reverse transcriptase is a multi-functional enzymewhich has the enzymatic activities of RNA-dependent DNApolymerase, DNA-dependent DNA polymerase and RNaseH. All of the three functions are essential to complete thereverse transcription. The polymerase activity of the HIV RT

*Address correspondence to this author at the Medivir AB, Lunastigen 7,SE-141 44 Huddinge, Sweden; Tel: +46 8 54683100; Fax: +46 8 54683199;E-mail: [email protected]

shares similar feature as most DNA polymerases, howeverwith a higher affinity to RNA as a template. Kinetic studieof the enzymatic reaction indicate that the DNA polymeriza

tion takes place in an ordered mechanism. The reaction isinitiated by the binding of the primer, in most cases thetRNALys , to the free RT, which is followed by the templatebinding to the binary complex. Then NTP binds subsequently to the complex. The multiple components complexundergoes a conformational change which is rate-limiting inthe polymerization, forming a tight binding complex thaenables the nucleophilic attack by the 3-hydroxyl of theprimer on the incoming phosphate of the dNTP [26-29]. TheRNase H activity catalyzes the degradation of the templateRNA in the RNA/DNA hybrid during the reverse transcription. Both endo- and exonuclease activities have been foundwith HIV RT. Two mechanisms of action have been ob-served with the RNase H activities in HIV RT, namely the

polymerase-dependent and polymerase independent. Theformer hydrolyzes the RNA template upon the elongation othe DNA chain and the latter removes the residual primaryRNA before the DNA strand transfer [30-32].

HIV-1 RT is a heterodimer, consisting of a p66 subuniwith 560 amino acids and a p51 subunit with 440 amino acids. The 2 subunits form a stable dimer which is essential forthe enzymatic activity. The overall structure of HIV-RT re-sembles a right hand as shown in Fig. 1 where the subdomains of p66 are designated as finger (residues 1-85), palm(residues 86-117 and 156-237) and thumb (residues 238-318regions. Besides, there are other subdomains as RNase H(residues 427-560) and connection (residues 319-426). The

conserved residues Asp185, Asp186 and Asp110 form thepolymerase active site in the palm domain [33]. Although thep51 subunit shares the same sequence as the N-terminal othe p66 subunit, the p51 subunit adapts a spatial arrangementotally different from the p66 subunit and the p55 subunit hano catalytic function as the corresponding Asp residues areburied in the structure. It is believed that p51 plays a role inmaintaining the overall structure of the RT. The p66 fingeand thumb domains are rather flexible and the nucleic acidtemplate and primer are bound in this cleft passing between


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102 Anti-Infective Agents in Medicinal Chemistry, 2008, Vol. 7, No. 2 Sahlberg and Zhou

the fingers and in front of the thumb domain. When dNTP isbound to the complex, the fingers bend towards the palmforming a catalytic pocket [34.35].

Due to the essential role of the HIV RT in the viral repli-cation, the inhibition of RT is regarded as one of the mostattractive targets in the anti-HIV chemotherapy. Based on the

structure and the mechanism, HIV RT inhibitors can be clas-sified as two groups, nucleoside or nucleotide reverse tran-scriptase inhibitors (NRTIs ) and non-nucleoside reversetranscriptase inhibitors (NNRTIs). The NRTIs, not discussedin this review, are competitive inhibitors to dNTP and needto be transformed through a phosphorylation process to theircorresponding nucleoside triphosphates or nucleotidediphosphates which then are incorporated into the growingviral DNA chain and subsequently to terminate the DNAelongation. They are normally active against both HIV-1 andHIV-2. The NNRTIs, on the other hand, are almost inactiveon HIV-2, need no metabolic activation and they bind to theenzyme independently from dNTP and template/ primer witha non-competitive kinetics to dNTP and are uncompetitive to

template/primer. Different aspects on the inhibition of HIV-1RT with NNRTIs are discussed below.

Inhibition of HIV RT by NNRTIs

The enzyme kinetic studies of the NNRTI in complexwith RT suggested an allosteric mechanism, which has beenconfirmed by several X-ray crystallography studies of theRT co-crystallized with different NNRTIs [33, 36-42]. Allknown NNRTIs bind to one specific pocket in p66 subunit,which is about 10 away from the catalytic site. The pocketis located in the palm subdomain and surrounded by severalstranded -sheets [34].The pocket is basically hydrophobic

in nature and lined with many aromatic and hydrophobicaliphatic residues as Leu100, Val106, Tyr181, Tyr188Phe227, Trp 229, Leu234 and Tyr232. It contains also somehydrophilic residues, Lys101, Lys103, Ser105, Asp192 andGlu224 (Fig. 2).

The NNRTI binding pocket (NNBP) exists only when an

inhibitor is bound to the enzyme [38, 43]. Upon binding ofan NNRTI to RT, the side chains of Tyr181 and Tyr188originally pointing to the hydrophobic center of the pocketare now pointing towards the direction of the catalytic siteThis conformation change creates the NNBP and this pockecan accommodate a space of about 620-720 3 which is approximately more than twice of the volume occupied bymost of the present NNRTIs [43]. The formation of the newpocket brings about the dislocation of -sheets where thecatalytic Asp residues sit, leading to a position shift oaround 2 for the catalytic residues [44]. The mouth foentering the pocket is flanked by Pro225 and Pro236 whichare located on flexible chains. The flexibility is crucial fopermitting the entrance of the NNRTI and subsequently clos

ing the mouth after the entrance of the NNRTI. A probablesolvent accessible area is located at the interface betweenp66 and p51 surrounded by Leu100, Lys101, Lys103 Val179and Tyr181 from p66 and Glu1138 and Thr1139 from p51.

The residues in the NNRTI binding pocket contribute at avaried degree to the affinity between the inhibitor and theenzyme. Some interactions are crucial and play an importanrole in a majority of the NNRTIs. The aromatic residues inRT, like, Tyr181, Tyr188 and Trp229, provide very important hydrophobic -interactions with -electron containingcomponents of the inhibitors. The inhibitors are also possible

Fig. (1). The overall structure of RT1

1From coordinates kindly supplied by Prof. Torsten Unge, Department of Cell and Molecular Biology, Uppsala university


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Development of Non-Nucleoside Reverse Transcriptase Inhibitors Anti-Infective Agents in Medicinal Chemistry, 2008, Vol. 7, No. 2 103

to increase their affinity to the enzyme through van de Waalsinteractions with various positions in RT i.e. Leu100,Val106, Val179 and Leu234. Interactions with Lys101,Lys103 and Glu1138 represent the possibility of catchingelectrostatic interactions. Many NNRTIs form H-bonds withthe enzyme through the side chain as well as backbone am-ides. Loss of some key interactions will significantly reducethe potency of the inhibitor. For example, Tyr181 plays an

important role in interacting with many NNRTIs and the factthat HIV-2 RT has a isoleucine in the 181 position partlyexplains the reason that many NNRTIs are inactive to HIV-2, although HIV-2 RT shares an similar overall folding asHIV-1 RT [45].

Three mechanism hypotheses for the inhibition of HIV-1RT by NNRTIs have been suggested and none of themshould be regarded as mutually exclusive. In fact, the natureof a particular inhibitor may dictate which mechanism isdominant.

The first mechanism suggests a disposition of the cata-lytic Asp residues. It has been shown by kinetic studies that aNNRTI present in RT has a relatively small effect on the

dNTP binding with an overall affinity reduction of around 10fold. Meanwhile the affinity of the primer/template duplex tothe enzyme is increased [26]. Hence, the major effect of theinhibition is believed from the blocking of the chemical step,which was confirmed by the analysis of the pre-steady stateburst of the DNA polymerization [44]. The structural studiesof the NNRTI bound enzyme clearly showed the dispositionof the catalytic residue, which can partly explain the block-age or low efficiency of the nucleotide transfer reaction.

The second mechanism suggests a hindrance of the coop-erative motion of the thumb/finger subdomains. The bindingof the NNRTI can also cause a long range distortion of the

RT structure. It was found that the p66 thumb subdomain irotated by 40 relative to the finger domains compared totheir relation in the NNRTI unbound enzyme. Associatedwith the hinge movement of the thumb, p66 connection andRNase H subdomains are also distorted from the normal position in unbound enzyme [36]. A dynamic study of RT sug-gested that the NNRTI binding not suppress the mobility othe finger, thumb and palm subdomains, but it will interfere

with the global hinge-bending movement that controls thecooperative motion of the subdomains [46].

The last mechanism suggestion is rather speculative, addressing the effect the NNRTI binding has on the interactionbetween the two subunits p66 and p51. The change of thedissociation free energy of the RT heterodimer has been observed when binding with a NNRTI, which may has an impact on the modulating of the enzymatic function, since theheterodimeric structure is essential for the RT activity[47,48].

The inhibition of HIV-1 RT is regarded as one of themost attractive targets in the chemotherapy againsHIV/AIDS due to its essential role in the viral replication

and NNRTIs have been proven to be important componentin HIV/AIDS combination therapy. Today three NNRTIhave been licensed for clinical use, namely nevirapineefavirenz and delavirdine (Fig. 3) and many structuraclasses are being explored.

NNRTIs IN CLINICAL USE

Nevirapine

Nevirapine (Viramune

) was developed and launched byBoehringer Ingelheim for the treatment of HIV infection[49,50]. As the first NNRTI to be introduced into HIV treat

Fig. (2). NNRTI pocket with inhibitor and relevant amino acid residues1

1Data from reference [40]


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ment, it dramatically changed the strategy of HIV therapy.Nevirapine has since been widely used in the combinationtherapy for HIV/AIDS in various treatment regimens.

Nevirapine is a relatively potent inhibitor of wt HIV-1with an IC50 of 39 nM in CEM T-lymphoblastoid cells and44 nM in macrophages, measured based on p24 antigen re-duction [49]. The IC95 value in MT-4 cells is about 400 nM.

The Ki value for wt HIV-1 in the RT enzyme assay is about200 nM. As a selective inhibitor of the viral polymerase,nevirapine has low inhibitory effect on cellular polymerasesand is noncytotoxic up to a very high concentration (CC50 =321000 nM). The substance is basically inactive againstHIV-2. Nevirapine shows a strong synergistic effect in com-bination with NRTIs, hence being widely used in the combi-nation therapy with anti-HIV nucleoside analogs. For exam-ples, in the plague assays the IC50 value for AZT is 10 nM,and for nevirapine 32 nM. In the combination experiment, 50% inhibition was achieved with 0.3 nM AZT and 10nM nevi-rapine giving a combination index of 0.24, which indicates astrong synergistic effect [51].

Nevirapine SAR has been extensively studied. Thedipyridodiazepinone structure is important for nevirapine.Changing either pyridine of the structure to pyrido[2,3-b][1,4]benzodiazepinone (compound 1) or pyrido[2,3-b][1,5]benzodiazepinone (compound 2) will reduce the activ-ity [52]. Although maintaining sufficient antiviral activity,the benzo compounds have much lower water solubility thannevirapine and are more susceptible to metabolic N-dealkylation in the N-11 position, which may have a nega-tive impact on their in vivo DMPK profiles compared tonevirapine. Though the diazepinthione modification (com-pound 3) is allowed and, in most cases, can enhance the anti-viral potency, the thiones are generally regarded as less at-tractive for development due to observations that many

thiolactams will be metabolized rapidly and that the they areoften less water soluble. The substitution on the lactam ni-trogen (N-5) is allowed when the 4-position is unsubstituted(compound 4). Increasing the size of the N-5 substituentswill reduce the activity. The substitution on N-11 is crucialfor potency (compound 5). Removal of the cyclopropylgroup abolishes the activity. The best activities are seen withsmall liphophilic groups, such as ethyl, propyl and cyclopro-pyl. Increasing the size of the substituent or introducing hy-drophilic group or electron-withdrawing group diminishesthe potency. Substitution on either pyridine rings as in com-pounds 6 and 7have been extensively explored. In general,

substituents in 2-position appear to have limited effect on theactivity, whereas substitution in the 3-position is unfavoredThe methyl substitution at 4-position results in good potency. However, the substitution at 7-, 8- or 9-position canaffect the potency negatively. Some of the 5-alkylated lactams substituted at 2-position with heterocycles (compound8) like pyrazol, pyridyl and thiazolyl give good potency

which confirms the great tolerability of the 2-position [53].The structural studies of nevirapine and its interaction

with RT has shed some light to the understanding of its SARand provided the useful information for the further modification of the structure [38,43,54,55]. Nevirapine adopts a butterfly-like conformation which is commonly seen with manyNNRTIs. The molecule is fold such that the angle of the twopyridine rings is about 120. Due to the electron delocalization, the amide bond moiety in the 7-membered ring adoptan almost planar conformation. The cyclopropyl substituenpoints up and away from the tricyclic system. One of thepyridine keeps a strong -stacking with Y181, Y188 andW229, while the other has interactions with K101, K103V106, V179, Y318 and even possibly with the main chain

atoms of H235 and P236.

Nevirapine has a very favorable DMPK profile. Thecompound is quickly absorbed after oral dosing, and the absolute oral bioavailability reaches as high as >90% in human[56]. At the same time, the compound has a long half lifeabout 25 hrs. When dosed at 200mg twice daily, it can reacha steady-state trough level of around 16 M which is around300 fold of the EC50 in cell culture. The protein binding leveof the compound in plasma is rather low compared to otherNNRTIs with a value of 60%. Nevirapine is mainly metabolized by CYP3A4 like other lipophilic NNRTIs. The hy-droxylated metabolites can further undergo glucuronidationbefore the excretion via urine. Interestingly, nevirapine i

also a strong inducer of CYP3A4. After 2-4 weeks dosingthe CYP catalyzed metabolism is found to be enhanced 1.5 2 fold. Therefore, the early recommended dose for nevirapine was 200mg once daily for the first two week and subsequently enhanced to 200mg twice daily [57]. However, therecent 2NN study has shown that it is possible to dose nevirapine QD together with 2 NRTIs with the same antiretrovi-ral efficacy as with the BID dose regimens thus simplifyingthe treatment and enhancing the patient compliance [58].

Nevirapine is a relatively safe drug although it is extremely important to follow-up the first 18 weeks of dosingto detect any life-threatening skin reactions such as Steven

Fig. (3). NNRTIs in clinical use

NN

HN

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S O

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OCl

O

Nevirapine Delavirdine Efavirenz

F3C


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Johnsons syndrome, toxic epidermal necrolysis and rash andalso life-threatening hepatotoxicity that can occur with theuse of nevirapine [59].

The very low genetic barrier seen with nevirapine has

had a great impact on the clinical use of nevirapine and is adriving force in the search of new NNRTIs. It was noticedvery early that the rate of resistance development was veryfast upon monotherapy with nevirapine [60]. The present useof nevirapine is of course as a part of combination therapyand has recently been reviewed [8]. The resistance develop-ment is still an debated issue as nevirapine is used in Africafor the prevention of HIV from mothers to infants and in arecent publication these aspects have been discussed and itwas concluded that resistance after single-dose nevirapineoccurs more readily than previously thought [61].

Delavirdine

Delavirdine (Rescriptor

) was originally developed byUpjohn and launched by Pharmacia&Upjohn for the combi-nation therapy against HIV infection [62]. The compound isnow marketed by Pfizer.

Delavirdine has a good activity against HIV-RT with aKi about 0.2 M for the wild type enzyme. In the cell cul-ture, the ED50 reaches low nanomolar level. Given the ex-tremely low cytotoxicity ( > 100 M), the compound has aselective index over 10000. However, the substance losessubstantially its potency against resistant strains with singlemutations such Y181C, K103N. which are commonly seen

with NNRTI treatment, as well as P236L which is specificfor delavirdine.

SAR of delavirdine has been thoroughly studied [63-68]Basically, the molecule can be regarded as three parts, i.e. a

left aromatic part, a central piperazine and a right aromaticpart (structure 9). In the central part, Y can be a carbonyl or amethylene group, with marginal effect on the activitiesHowever, varying the length of the spacer Y will significantly decrease the inhibition of RT. The best activity is seenwith a 1 atom spacer. The right hand pyridine structure icrucial for the potency. Replacing it with phenyl ring or sub-stituted phenyl ring will abolish the activity. Similarlypyrimidine replacement will also destroy the potency (structure 10). The isopropylamino substituent on the pyridine isimportant (compound 11), however, not essential. Smallealkylamino displacement is tolerated. Many aromatic ringsincluding substituted phenyl, aromatic heterocycles andfused aromatics, were explored as the left hand aromatic

part. Although several aromatic rings maintain good activities, such as thienyl, benzofuryl and naphthyl, indole rings ingeneral have a superior antiviral potency and furthermore areduced inhibitory effect to the cellular DNA polymerasesIn particular, the indoles being connected through 2-position(compound 12). Various substitutions on indoles were evaluated, 5-substituted analog were selected due to its goodmetabolic stability and relatively good potency (compound13). A large variation of substituents is allowed in the 5position; for example, halogen, alkoxy give a similar activityas the methylsulfonamido group. Alkylaminopiperidinewere studied as an alternative to the central piperazine struc

Fig. (4). Nevirapine SAR.

NN

HN

N

O

NN

HN

O

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HN

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HN

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S

NN

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Nevirapine

1

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45

6 7

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1011

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Het


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ture. While the potency against wild type RT is a bit lowerthan delavirdine, compound 14 is slightly more activeagainst wild type HIV in cell cultures and substantially moreactive against mutant RT such as Y181C or P236L [69].

Delavirdine occupies the same NNRTI binding pocket asother NNRTIs however with a very different binding modeas delavirdine is bulkier than the other NNRTI with a vol-ume of about 380 3 opposed to 230- 290 3 for the others.The compound has also an overall shape which differs dra-matically from other NNRTIs. The complex between de-lavirdine and RT is stabilized in a unique way; when bond,delavirdine extends beyond the usual pocket and projects

into the solvent [70]. A few hypothesis were suggested as forthe entry of delavirdine into the NNRTI BP [33, 37]. Thedelavirdine binding pocket is connected to the solventthrough a channel between well-ordered -sheets and ahighly mobile flap, and the entry of delavirdine is by meansof this channel. Delavirdine also interacts with regions ofNNRTI BP inaccessible to other NNRTIs. For example, thepiperazine ring positions very close to V106 and the indolering forms a strong hydrophobic interaction with P236 withseven inter-ring atom distances being less than 3.6 . Be-sides, the carbonyl oxygen in the central spacer is H-bondedto the main chain of K103.

Delavirdine is well absorbed in vivo. In human it cangenerally reach over 50% absolute oral bioavailability and insome patients even over 85% [71]. Delavirdine has nonlinear pharmacokinetics, showing 2 compartments and a substantial individual variation [72]. Thus, delavirdine was initially developed with an original dosage of 100mg TID anddue to the above reasons the recommended dosage was fi-nally set at 400 mg TID. Following the absorption, delavirdine distributes across cell membranes with a steadystate volume of distribution of around 0.8 L/Kg. The com-pound has a high protein binding (98%). The plasma half lifeis about 7-21 hours. Delavirdine undergoes extensive N

dealkylation, forming N-deisopropyl delavirdine as the majometabolite [73]. The main enzyme responsible for the metabolism of delavirdine is CYP 3A4. Besides, CYP2D6, 2C9and 2C19 are also important in the metabolism of delavirdine. Delavirdine is also an inhibitor of CYP3A4 [74which could potentially enhance the plasma levels of certainPIs that are metabolized by CYPA4 when co-administeredwith delavirdine. The absorption of delavirdine is much affected by the gastric pH and its solubility can decrease 200fold when pH is increased from pH 2 to pH 7.5.

The most common adverse effect associated with delavirdine is different degrees of skin rash. Hepatic transa

Fig. (5). Delavirdine SAR.

HN

NH

S O

N

N

N

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O

HN

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S O

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minase activity elevation has been reported. [75]. De-lavirdine treatment can develop resistance quite rapidly andthe compound loses substantially the efficacy against mutantstrains of HIV virus. With a relatively high and inconvenientdose regimens involving 400 mg/3 times a day, relativelylow potency and an unfavorable resistance profile, the com-pound is not as widely clinically used as the other twolaunched NNRTIs.

Efavirenz

Efavirenz (Sustiva) was developed by DuPont and laterby BMS for the treatment of HIV infection in combinationwith other anti-HIV agents [76]. The product was first ap-proved by FDA in 1998 for use in the triple combinations asan alternative to protease inhibitors. Due to the high potencyagainst wild type and some mutant strains, excellent PK pro-file, the compound is regarded as the leading prescribedNNRTI as a safe and efficacious component of earlyHAART regimens.

Efavirenz is a highly potent inhibitor of HIV RT with aKi of 2.9 nM against wild type enzyme and an ED50 of 1.5

nM in MT-4 cells. The compound is not cytotoxic and has aCC50 of about 80M which gives a selective index about80000. Despite with certain loss, it maintains good activitiesagainst many single mutants commonly occurred in HIVtherapy such as the L100I, K101E, V106A, V179D andY181C with Ki values ranging 7-18 nM and ED50 valuesranging 3-100 nM. However, it loses potency against doubleor triple mutants. For example, the ED50 values againstK101D+K103N, L100I+K101D, K103N+Y181C andL100I+K103N were reported to be 1.5 M, 1.5 M, 0.4 Mand 25 M, respectively [77].

Efavirenz was found as the result of the optimizationwork starting from the lead structure 15 [76]. The thioureastructure has a good activity against HIV-1 RT, however,due to the metabolic instability and the potential safety risk,it was replaced by the urea scaffold with 4,4-dialkyl substitu-tion, which maintained good potency and stability as exem-plified by structure 16. The N-3 methyl group has limitedeffect on the potency. The choice of the alkyl group in the 4-position groups is important for rendering the potency aswell as oral bioavailability. Compound 17 is highly potent incell culture with an ED50 of 12 nM [78]. Interestingly, thestereochemistry is crucial for the antiviral activity. The com-pound with (R) configuration in the 4-position (compound18) is basically inactive with an ED50 of 9.3 M, whichcould be explained by an unfavorable binding to RT. Replac-ing the dihydroquinazolinone scaffold with a benzoxazinone

scaffold led to the a further enhancement of potency and theimprovement of PK profile, cf. structure 19 [79]. The R-groups in 19 were explored in combination with the substitu-ent in 6-position. When R is a small alkyl groups, it oftengives better activity. Halogen substitution on 6-position issuperior to electron donating groups such as amino, mono ordi-alkylamino, alkoxy, alkyl or phenyl groups. Multi substi-tution pattern on the benzene ring were also studied [80].Introducing more halogens in 7 or 8-position (compound 20)is tolerable, though with about 10 fold reduction of activity.However, introducing halogen substitution on both 7 and 8positions, forming a 6,7,8-trihalo compound, will totally

abolish the activity. Interestingly, on the other hand, the in-troduction of difluoro to the 5,6 position of the dihydro-quinazoline scaffold resulted in compounds more potent onthe K103N mutation exemplified by DPC 963 (Fig. 6) adescribed by Corbett et al [81]. Reduction of the acetylenicbond to a cis-olefin and keeping the 6-chloro substituenresulted in a compound named DPC 083 that has both a highantiviral activity and a promising PK profile such as lowe

protein binding than efavirenz and a long half lives whendosed orally to chimpanzees. DPC 083 went into clinicatrials but the development of the compound has most likelybeen discontinued.

Similar to most NNRTIs, efavirenz binds to RT throughstrong hydrophobic interactions [41, 82]. The cyclopropylpropynyl group points to a sub-pocket surrounded by aromatic side chains of Y181, Y188, W229 and F227, where theleft pyridine ring of nevirapine binds. The benzoxazinonering is situated between the side chains of L100 and V106 ina sandwich shape. The edge of the benzoxazinone has con-tact with the residues Y318 and V179. A prominent nonhydrophobic contact is the H-bonding formed between theNH in benzoxazinone and the main chain carbonyl oxygen oK101. The interaction of efavirenz with the mutant RT hasbeen a subject of great interest and the structural informationshed some light to the understanding of the mechanism othe efavirenz resistance. K103N is one of the major mutations efavirenz and other NNRTIs encounter. The structureof efavirenz with K103N mutant clearly shows that the muta-tion causes a significant conformational change within thebinding pocket. The bigger asparagine side chain pusheefavirenz deeper into the pocket, which in turn forces theside chain of Y181 flip to opposite direction and its OH idisplaced for more than 8 . This affects the position oE1138 and K101. The displacement of efavirenz itself iabout 0.5 for the carbonyl oxygen and 0.7 for the cyclopropyl group, which is sufficient to alter some contacts between efavirenz and the enzyme. However, efavirenz losesthe activity against single mutants much less than nevirapinein general. A hypothesis is forwarded from the structurastudy to explain the phenomenon. The overall size oefavirenz in relation to its binding size may determine itsability to adapt a mutation. A smaller inhibitor can repositionitself within the pocket whereas nevirapines bulky ring system remains relatively rigid and no such repositioning ispossible [81].

The PK of efavirenz has been extensively studied. Astudy in rats and monkeys indicated a good oral bioavailabityof 16 and 42%, respectively and long half life especially inmonkeys was observed [83]. The long half life is an impor-

tant property which potentially enabling a QD regimen. Further studies have confirmed this and the clinical dose foefavirenz is 600 mg QD. The PK properties are fully described in the prescribing information for the drug [84]Efavirenz is a highly lipophilic compound with a log P value> 4 and an aqueous solubility


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The metabolism is primarily catalyzed by CYP3A4 andCYP2B6. At the same time, efavirenz has certain degree ofinhibitory effect on CYP3A4, CYP2C9 and CYP2C19,which may lead to some drug-drug interaction. The metabo-lites are excreted through bile and urine, with less than 1%appearing as unchanged drug in urine [84].

The clinical efficacy of efavirenz is well documented forexample in the 2NN study which involved 1216 treatmentnave patients [58]. Treatment failure occurred in 151 pa-tients (38%) of 400 assigned to efavirenz 600 mg QD incombination with stavudine and lamivudine which was

somewhat less than the group receiving nevirapine QD orBID. There is a possibility that the efficacy of efavirenz ismore sustainable than of nevirapine BID. The safety profilewas better for efavirenz and more importantly the arm withboth efavirenz and nevirapine gave most treatment failuresand adverse effects showing that in contrast to NRTIs andPIs combinations of NNRTIs should be avoided. The advan-tage of using efavirenz in salvage therapy is not as well es-tablished. In a recent study the use of efavirenz as a HAARTcomponent was compared in treatment nave patients andpatients with treatment failures from different PI-containingregimens was compared [86]. The result indicated clearly

that the efavirenz initial triple HAART therapy led to a betteroutcome than the use of efavirenz in the salvage group. Thimight be explained by the broad spread of viral resistant viruses and cross resistance over time. Actually, it was alsosomewhat surprising as it occurred in patients never takenNNRTI drugs. This observation seems to point to a relativelylow genetic barrier of efavirenz, which also has been shownin several in vitro studies. The clinical use of efavirenz hasrecently been reviewed [10].

The major adverse reactions reported with the efavirenztreatment are CNS related, skin rash and liver and transa

minase activity enhancement [84]. Skin rash is a commonadverse effect for all NNRTIs whereas the CNS disturbanceincluding nightmares are specific for efavirenz. These adverse effects together with a relatively low genetic barriermake efavirenz not to be an ideal drug although efavirenz icurrently the mostly prescribed NNRTI and is recommendedfor the first-line therapy. There is certainly room for newNNRTIs with properties such as high potency on wt viruand resistant virus, a high genetic barrier, DMPK propertieenabling QD dosing and an improved safety profile. Thenext section will deal with new NNRTI compounds undedevelopment.

Fig. (6). Efavirenz SAR.

NH

OCl

F3C

ONH

NCl

S

OEt

NH

NCl

O

R1 R2

NH

NHCl

O

N

NH

NHCl

O

N

NH

OCl

F3C

O

R

NH

OCl

F3C

O

XNH

NHCl

F3C

ONH

NHF

F3C

O

F

Efavirenz15

16

17 1819

20 DPC 083DPC 963

4

8

6

7

5


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NNRTIs IN DEVELOPMENT

Nevirapine Analogs

The low genetic barrier of nevirapine has initiated asearch for compounds more active on resistant forms ofHIV-1. Nevirapine analogs with an arylalkyl tail on the 3-ring system were explored, leading to an enhancement of thepotency against certain HIV-1 mutants. Compound 21 in Fig.

7 is an example of such a compound with a much improvedpotency on the K103N+Y181C double mutation. The com-pound with an un-oxidized sulfur has, not unexpectedly, alow metabolic stability and was not explored further [87]. Inan another paper, BI scientists described a series of com-pounds with carboxylic acids in the end of the tail [88].Compound 22 was high lightened as showing a good meta-bolic stability but the compound had problem to cope withthe single mutations Y188L and V106A. Further develop-ment led to BILR 355, a compound with a N-oxide contain-ing tail [89]. The compound has an excellent antiviral profilewith IC50 values below 6 nM in culture against many mu-tants including K103N and Y181C and the 103+181 doublemutation, However, the compound has only moderate activi-

ties against the single mutations Y188L and V106A and thedouble mutations K103N+V106A and Y181C+G190A.BILR 355 has a shift of about 2 fold upon addition of 50%human serum (HS) to the assay. The metabolic stability ofthe compound is substantially increased in rats when co-dosing with ritonavir indicating that the compound is me-tabolized by CYP3A4. BILR 355 is now in phase II clinicaltrials and needs to be co-dosed with ritonavir for achievingan acceptable AUC and half life.

Fig. (7). Nevirapine analogs.

Diaryl Pyrimidine Series (DAPY)

The properties and multidisciplinary efforts leading toTMC278, which is the latest compound in the diaryl

pyrimidine (DAPY) series of compounds from Janssen/Tibotec, are described in a recent paper [90]. The histori-cal background from TIBO to the present DAPY compoundsis also nicely summarized in the paper. The structures of thethree major DAPY compounds, TMC120, TMC125 andTMC278 are depicted in Fig. 8.

Fig. (8). Diaryl pyrimidines (DAPY).

TMC120

TMC120 is now developed as a vaginal microbicide

despite promising clinical efficacy seen with TMC120 as anoral drug as reported 2001 [91]. Tibotec out-licensed 2004the compound to the international society for microbiocide(IPM) to be used as a vaginal microbicide for preventing theHIV infection in females [92]. A recent publication described the delivery of TMC120 from silicone elastomevaginal rings in amounts sufficient to inhibit the HIV-1 replication [93].

TMC125

The initial SAR and anti-HIV-1 activities for the DAPYcompounds that led to TMC125 were first reported in a paper from 2001 [94]. TMC125 was the most potent compound

in the series with an impressive cell culture efficacy both onwt and on resistant forms of HIV-1 with IC50 values below 7nM for wt HIV-1 and HIV-1 virus with the mutations L100I, K103N, Y181C, Y188L, L100I+K103N andK103N+Y181C. The binding properties of TMC125 andearlier compounds in RT were assessed by the studies on x-ray crystallography of the complexes between RT and inhibitors and molecular modeling [95]. Surprisingly, there wasnot possible to get any structural information from the complex of wt-RT and TMC125 as this complex resolved aabout 7 but it was possible to get a good resolution of 2.9 on the K103N-RT and TMC125 complex. These com

NN

HN

N

O

S

N

NN

N

N

O

O

COOH

NN

N

N

O

O

N+

O-

22

BILR-355

21

N

N NHHN

CN

N

N NHO

CN

Br

CN

NH2

N

N NHHN

CN

CN

TMC120 (dapivirine) TMC125 (etravirine)

TMC278 (rilpivirine)


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pounds bind in horseshoe fashion and a suggestion whyTMC125 and analogs are still active on various resistantforms of HIV-1 was given. The DAPY compounds, beingquite flexible, have according the authors 2 ways to over-come the usual negatively effect mutations have on the af-finities of RT inhibitors in the NNRTI binding pocket(NNIBP) (i) by torsional flexibility (wiggling) in a givenbinding mode where the inhibitors can adapt numerous con-

formers enhancing the binding to mutant HIV-1 and (ii) thereis a possibility of repositioning and reorientation (jiggling)of the compounds within the allosteric pocket.

The original SAR work was followed up with a publica-tion describing the antiviral efficacy for TMC125 in variousmutants and also in comparison with nevirapine, delavirdineand efavirenz in different virus strains in MT4 and PBMCcells, in patient isolates with multiple mutations and in arange of HIV-1 subtypes. The metabolic stability and effecton addition of human proteins in the cell culture assays werealso studied [96]. As previously discussed, TMC125 has avery good antiviral efficacy on HIV-1 and also some modesteffect on HIV-2 with an IC50 of 3.5 M. In the NNRTI resis-tant strains (1081 samples with 10-fold IC50 change com-pared to wt for at least one of the clinically used NNRTIs)TMC125 inhibited 77% of the strains with an IC50 below 10nM whereas the figure for efavirenz was only 23%. Thisclearly indicates that TMC125 can be useful in patients in-fected with NNRTI resistant virus. TMC125 has a good sta-bility in human microsomes with 85% compound remainingafter a 2 h incubation. Despite a very high proteinbinding of>99%, TMC125 showed no or a very little fold shift of effi-cacy in cell culture when different human proteins such as1-acid glycoprotein (AAA), human serum albumin (HSA)and human serum (HS) were added. Nevirapine showed asimilar pattern whereas efavirenz was negatively affected bya factor of 20 upon the addition of 50% HS. The genetic bar-rier, as an important property for a NNRTI, has recently beeninvestigated in vitro for TMC125 [97]. The rate of resistancedevelopment i.e. breakthrough of mutants were studied bysequential passage experiments using 2 different approaches,a) a low multiplicity of infection (MOI) using increasingconcentrations of the inhibitor or b) using virus with highMOI and fixed concentrations of the studied compounds.The studies were conducted both with wt and resistant virus.The results indicated that multiple mutations were needed toget resistant virus insensitive to TMC125 and the time toresistance was considerably longer than that for nevirapineand efavirenz. Furthermore, when using the high MOI and afixed concentration of TMC125 of 1.0 M, no virus break-through occurred after the studied 30 days. The effect of 40

nM TMC125 was equivalent to 1.0 M efavirenz i.e 100%virus breakthrough at about day 10. The TMC125 selectionexperiments yielded known NNRTI mutations such as L10I,Y1811C, and G190E as well as novel ones like V179I andV179F. However, those in vitro selected mutations may notnecessarily lead to same outcome in vivo. For example, ca-pravirine gave V10A/F227L in vitro but studies in in vivoyielded mutations in positions 101, 108, 190 and/or 188.This observation is further illustrated by the fact that in vitroexperiments with efavirenz give not the K103N mutationthat is the main mutation seen in vivo.

The phase IIb trials of this compound are completed andseveral phase III studies are under way. A phase II studywith TMC125 on protease-inhibitor (PI) nave patients withNNRTI resistant virus was discontinued due to inferiority tothe PI containing controls and the reason for this is not cleafor the moment [98]. According Tibotec, this study has noeffect on the phase III registration studies for TMC125which are substantially different from the discontinued study

in terms of trial design, patient population, formulation andbackground regimen. Data from 3 short-term (7 days), phaseIIa clinical studies, dosing 900 mg BID of TMC125 formulated in PEG 4000, have been published. In the first studyTMC125 replaced nevirapine or efavirenz components in atriple drug therapy consisting of NRTIs and 1 NNRTI. Sixteen male patients were included with an mean viral load of4.2 log10 RNA copies/ml, among whom 81% were nevirapine experienced and 19% efavirenz experienced. The mediandecrease in viral load in the study was 0.89 log10 RNA copies/ml [99]. The second study compared the results frommonotherapy with TMC125 in antiretroviral therapy (ARTnave patients with a 5-drug regimen containing zidovudinelamivudine, abacavir, indinivair and nevirapine also in ART

nave patients. Twelve patients with a median viral load of4.2 log10 copies /ml received TMC125 and 11 patients witha median viral load of 4.8 log10 copies/ml received the 5drug regimen. The results indicated similar decrease of viraload for the 2 regimens i.e. at day seven the median viralload was 1.92 for the TMC125 regimen and 1.76 for the 5drug regimen [100]. In the third study, 19 ART nave pa-tients received TMC125 in a double-blind placebo controlledphase IIa clinical trial. In the TMC125 group the mean decrease after 7 days was 1.99 compared to 0.06 for the pla-cebo group [101]. All these 3 clinical studies indicate thaTMC125 dosed 900 mg twice a day has a good anti-HIVeffect.

TMC278

As previously mentioned, the multidisciplinary effortleading to TMC278 is described [90]. The SAR and synthesis of this compound and other related compounds are de-scribed in another paper [102]. TMC278 has a very highpotency in the cell culture system studied with IC50 below 8nM for all mutants tested including the double mutantK103N+Y181C and L100I+K103N. The compound is relatively easy to prepare in 8 steps but has a risk of isomerization of the E-form to the Z-isomer in day-light and in a weakacid solution. The rationale behind the series was to utilizethe interactions with the conserved W229 region. The mospotent structures found were , -unsaturated nitriles which

finally led to TMC278. The choice of this compound as theclinical candidate is somewhat surprisingly as it contains apotentially risky position which can react with various typesof nucleophiles. This might be reflected by the relatively lownon-toxic effect dose (NOEL) of 5 mg/kg reported for safetystudies in dogs [90].

The compound is in phase IIb clinical studies and theoutcome of a short study with TMC278 is recently published[103]. The study was a double-blind placebo controlled 7day study using once daily dosing of 25, 50, 100 or 250 mgTMC278 to 47 ART nave HIV infected patients. The resultshowed, independent of the dose, a median drop of viral load


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in the TMC278 treated group of about 1.2 log10 copies/mlcompared to 0.002 log10 copies /ml gained in the placebogroup. The plasma concentrations in all subjects were, at alltime points and for all doses, exceeding the targeted concen-tration of 13.5 ng/ml.

Imidazoles

Capravirine (S-1153), see Fig. 9, was originally devel-oped by Shionogi, but later licensed and further developedby Agouron/Pfizer. Antiviral key-data of capravirine on wtHIV-1 and various mutant forms of HIV-1 has been reported[104]. The compound has IC50 values below 2 ng/ml againstwt HIV-1 on various strains and in different cells and IC50values ranging 0.3 to 7 ng/ml for many mutants includingY181C and K103N in MT-4 cells. Furthermore, S-1153 hasIC50 values below 10 ng/ml for various clinical isolates andis considerably more potent than AZT on these isolates. Thecompound shows a substantially slower rate of resistancedevelopment than nevirapine when passaged in an in vitrosystem. It took more than 9 passages to generate resistantvirus (IC50 =740 ng/ml) with the double mutationV106A+F227L compared to 2-3 passages for nevirapine.Phase I clinical studies started in 1997 and some clinical datahave been published [105]. The results indicate a good anti-HIV-1 effect when dosing 25 mg/kg three times a day (TID)with a decrease of HIV-1 viral load with 1.34 log10 cop-ies/mL. The compound has a relatively short half life in hu-mans which has been further determined to be about 1.8 hupon a single dose of 1400 mg. The half life is increased to8.3 h when co-dosed with ritonavir. The compound is exten-sively metabolized according to a very complicated meta-bolic pattern [106]. The compound was in phase II/III clini-cal studies for many years and the development finally dis-continued as the combination therapy with capravirine as acomponent could not show a better effect than the standard

triple drug regimens [107].

Fig. (9). Imidazoles.

Gilead has filed an extensive patent on various NNRTIslinked phosphonates in order to make NNRTI prodrugs andthereby potentially enhance the half-life of the parent compound [108]. One of the specifically claimed compound wathe prodrug of capravirine (23), depicted in Fig. 9. Howeverno biological data supporting a longer half-life in vivo wagiven in the patent.

The Rega institute has patented various imidazoles re

lated to capravirine [109]. One example of a specificallyclaimed compound is structure 24 in Fig. 9 with a modesIC50 value of 0.2 g/ml in MT-4 cells.

Benzophenones

Chan et al reported a group of highly potent benzophenone derivatives active both on wt virus and various HIV-1mutants [110]. The compounds, GW4511, GW4751 andGW3011, cf. Fig. 10, showed excellent effect on wt and 16different single and double mutations, with IC50 values below or equal 2 nM for wt virus and less that 10 nM for mutant virus including the clinically important Y181C andK103N mutations. The work was followed up with a suc

cessful SAR work leading to the clinical candidateGW678248 which has excellent anti-HIV-1 effect with IC5values of 0.5 nM on wt virus, 1 nM on the K103N mutantand 0.7 nM on Y181C mutant in cell culture systems usingMT4 cells [111]. A prodrug, GW695634, was made in ordeto increase the solubility and thereby the oral availability oGW678248 (Fig. 10). This prodrug was selected for clinicastudies. The antiviral properties, protein binding shift andcytotoxicity of GW678248 were further studied and reportedby Boone et al [112]. The excellent antiviral effect was confirmed and compared with nevirapine and efavirenz. Theonly mutants that gave IC50 values above 21 nM were singleY188L and triple V106I, E1138K and P236L which wasformed from serial passages of wt virus. The compound ha

a protein binding shift of about 7 and a favorable selectiveindex of over 2500 when comparing the cytotoxic effect withthe antiviral effect on wt, Y181C and K103N HIV-1.

Phase IIb studies with GW695634 showed good antiviraeffect when dosed 100, 200, 300 or 400 mg BID as mono-therapy [113]. However, the development of the compoundhas been discontinued due to safety issues related to rash andliver metabolic enzymes [6].

PETT Series

The phenyl ethyl thiourea thiazole (PETT) series oNNRTIs originating from a collaboration between Mediviand Lilly which resulted in the clinical candidate trovidine

with a IC50 value of 20 nM on wt HIV-1 when tested in MT4 cells [114]. Fig. 11 shows the structures of some representative PETT compounds. The emerging of the NNRTI resistance issues resulted in search for compounds active on resistant forms of HIV-1. The introduction of conformationallyrestricted cyclopropanes in the PETT series was beneficiafor the activity on resistant forms of HIV-1 and the change tourea compounds was also beneficial for lowering the relatively high protein binding shift seen in cell culture with thethiourea compounds. The stereochemistry was also studiedas well as extensive X-ray studies of the complexes betweenthe inhibitors and both wt and resistant HIV-1. This work

S

N

N

N

O NH2

O

Cl

Cl

N

N

HN

Cl

Cl

S

N

N

N

O NH

O

Cl

Cl

P

O

OO

Capravirine (S-1153)

24

23


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resulted in MIV-150 and analogous compounds [40]. Theinteraction between wt- RT and MIV-150 is very well char-acterized as shown in Fig. 12. Very important is the internalhydrogen bonding net-work giving a derivative with optimalconfiguration to be bound to the enzyme. Interestingly, thetwo cis-enantiomers have similar potency which can benicely explained by x-ray studies. A separate x-ray studywith MSC-194, cf. Fig. 11, efavirenz and PNU14721 (aNNTRI from Pharmacia-Upjohn) on K103N mutant enzymein comparison with wt structure of efavirenz showed that the

K103N substitution gave no major changes in the positioningof the inhibitors in the NNIBP. It was concluded that theinhibitors bind in a conservative mode to K103N mutant andthat the changes in affinity was attributed to changes in thechemical environment close to the mutated 103 position [41].

MIV-150 was taken into clinical studies by Chiron andan optimized the process route was recently published [115]Due to the relatively low oral availability, MIV-150 is nowbeing developed as an vaginal microbicide by the PopulationCouncil [116].

The focus on the K103N mutation resulted in the com-pound MIV-160 (Fig. 11) described in a patent [117]. TheIC50 values, when tested in Medivirs cell culture system onwt and K103N, were 2 and 24 nM, respectively. MIV-160

was also tested at Virco and the IC50 values for wt andK103N were 0.3 and 0.7 nM, respectively, showing the welknown fact that it is very important to compare compoundin the same assay. MIV-160 has a fold decrease of about 5on the addition of 40% HS to the cell culture assay. In con-trast to MIV-150 and analogs, it is only the (-)-enantiomer o

Fig. (10). Benzophenones.

Fig. (11). PETT series (Medivir).

Cl

O

Cl

O

O

HN

SNC

NH2

O

O

Cl

O

Cl

O

O

HN

SNC

NH

O

OO

R1

O

Cl

O

O

HN

SR

NH2

O

O

F

O

Cl

O

O

HN

OF3C S

O

O NH2

GW678248 GW695634

GW3011R=CF3, R1=F ; GW4511R=H, R1=CN; GW4751

O

F

F

H

H

H

NH

NH

O

N

CN

N NH

NH

S

N

Br

NH

NH

O

N

CNF

OH

O

NH

NH

S

N

CNO

OH

O

Trovirdine

MSC-194

MIV-150

MIV-160


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the two possible endo (cis) tricyclic compounds that is bear-ing the anti-HIV activity.

Uckun and co-workers have further explored the non-cyclopropyl thiourea compounds and were able to patentsome limited analogs whereof compound 25 in Fig. 13 wasreported as the most potent one with a subnanomolar potencyon wt HIV-1 in a cell culture assay using PBMC cells [118].

Some other compounds reported by Parker Hughes are alsoshown in Fig. 13. They extended the known positive effectof a methyl group in the benzylic position [114] and studiedthe stereochemistry of alkyl substitution at this benzylic po-sition [119]. The results indicate that R-stereochemistry isoptimal as exemplified for compound 26 in Fig. 13 which isabout 10 times as active as the S-enantiomer.

Fig. (13). PETT series (Parker Hughes Institute).

Interestingly, another PETT derivative i.e compoundPHI-236 in Fig. 13, was recently reported to have good ef-fect in preventing HIV infection when used vaginally in amouse model [120]. This further provides an extra support tothe on-going clinical studies with MIV-150 and TMC120 asvaginal microbicides against HIV transfection.

Pyrimidindiones

HEPT was along with TIBO the first reported NNRTI. Ihas its origin from the NRTIs and has a modest IC50 value o5 M on wt HIV-1 [121]. Optimization of the HEPT originastructure led to MKC-442 which also is named emivirine(Fig. 14). The compound has an IC50 of about 15 nM againswt virus in MT4 cells [122]. The safety and pharmacokinet

ics of the compound have been reported [123]. Emivirinewas licensed to Triangle Pharmaceuticals and entered clinical trials but returned back to Mitsubishi in 2004 when it hadreached phase II. The present status of the compound is un-known.

SJ-3366 is another HEPT related compound, discoveredand developed by Sam Jin Pharmaceuticals. The structure isshown in Fig. 14. The anti-viral characteristics of this compound has been described [124]. The compound has a duaeffect on HIV. Besides being a allosteric inhibitor of RT, ialso inhibits the entry of the HIV virus into the target cellsHence, on the contrary to most NNRTIs, the compound isalso active on HIV-2 with an IC50 value of 170 nM in aCEM-SS cell system. The compound has high potency in arange of HIV-1 isolates and cells with IC50 values between0.9 and 10 nM. The addition of serum proteins gave basi-cally no effect on the potency of the compound except forthe addition of HSA+ AAA which caused a protein bindingshift of 10. The major drawback with the compound seems tobe the very low potency on K103N and Y188C mutant viruand an unoptimal effect on Y181C resistant virus with thefold resistance of >83, 33 and 5, respectively. SJ-3366 warecently out-licensed to ImQuest Pharmaceuticals and is renamed IQP-0410 [125]. An IND application was expected tobe filed 2006 according the homepage of the company.

Fig. (12). Interactions MIV-150 and wt HIV-1 RT.

N

N

N NH

O

OH

F

H

O

Provides angle

Pi-interaction with Phe227

EWG, preferably slightly bigger

H-bonding to Lys101 backbone

Interactions with Glu1138?

EWG, interaction with the "edge" of Phe227

Close-packingwith Trp229

Pi-stacking withTyr181, 188

H-bonding network

N NH

NH

S

N

Br

NH

NH

S

N

Br

O

O

NH

NH

S

N

Br

25

PHI-236

26


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Fig. (14). Pyrimidindiones.

Compound 27 in Fig. 14 was designed by the aid of mo-lecular modeling, and is a conformationally restricted NH-DABO derivative that was recently published by Italian sci-entists. The compound has a modest IC50 on wt virus of 30nM and shows certain inhibitory effect (IC50 =160 nM) onthe nevirapine mutant, Y181C [126].

Triazoles

In 2006 two groups reported independently, about newseries of NNRTIs using a triazole as the scaffold. In the firstpaper, compound 28 in Fig. 15 and analogs thereof weredescribed [127]. This compound is one of the most activecompounds in this sulfanyltriazoles series and shows singledigit nM potent on wt HIV-1 and a relatively low decrease(24 fold) on the double mutant K103N+Y181C. However, itloses potency heavily against the Y188L single mutant (3500fold). A second publication deals with compounds with the

Fig. (15). Triazoles.

same scaffold although somewhat more advanced and decorated leading to compounds with increased potency on mutant forms of HIV-1 and especially on the Y188L mutationThis series of compounds is exemplified with compound 29which shows only 4 fold loss on Y188L. This mutant causemuch more problems to these triazole types of compoundsthan the K103+Y181C double mutation [128]. The seriewere originally developed by Valeant and one compound

from the series was likely chosen as a clinical candidate(AR-806). Late 2006 this compound was acquired by Ardeabiosciences and a phase I study is planned for Q2 2007[129].

CONCLUSIONS

The use of NNRTIs as an important part of successfucombination therapy in HIV/AIDS patients have been welestablished and widely accepted. The drugs in clinical usenevirapine, delavirdine and efavirenz, are relatively old andhave their drawbacks. Nevirapine has a very low geneticbarrier and certain adverse effects associated with liver toxicity; delavirdine has also a low genetic barrier in addition toits unfavored TID dosing and efavirenz an insufficient genetic barrier and frequent CNS side effects. All three compounds have problems with different degrees of skin rashNevirapine and efavirenz have PK properties such as theycan be dosed once daily which is an important property for aHIV/AIDS drug to be competitive. However, the draw-backwith the existing compounds create of course a great needfor new NNRTI drugs.

Although the enormous progress that has been made inthe NNRTI field in recent years, especially in terms of theantiviral potency, the NNRTI clinical pipeline seems not tobe that impressive as hoped. The only compounds remainingin phase II/III trials seem to be the TMC125 and TMC278from Tibotec. These compounds have excellent potency on

HIV-1 and mutants thereof and probably a high genetic barrier which will cause a slow resistance development. Also tobe successful the compounds need to have satisfactory safetyproperties and very importantly to be dosed once daily (QD)This pharmacokinetic property is not obvious for TMC125but it seems that TMC278 can be dosed once daily. Regarding safety issues, TMC278 contains a potential reactivecomponent that could jeopardize its further development.

The need for new NNRTIs to be combined with otherdrugs, NRTis and/or PIs is still high. Reviewing the experi-ences from the NNRTI development and the clinical resultsit point to several key properties an successful NNRTI mayneed to have, namely excellent potency on wt virus and

clinically important mutations, high genetic barrier for resistance development, QD dosing, minimum adverse effectgood combination with other anti-HIV agents like NRTIsPIs or fusion inhibitors and satisfactory DMPK and safetyprofiles are much motivated and can be highly rewardingTime will show if any of the compounds described in thireview will fulfill these criteria.

ACKNOWLEDGEMENTS

The authors would like to thank Katarina Jansson formaking the Figures 1 and 2.

HN

N

O

SO

O

HO

HN

N

O

O

O

HN

N

O

O

O

HN

N

O

HN

F F

HEPT MKC-442 (Emivirine)

SJ-336627

S

O

HN

NO2NN

N

S

O

HN

NN

NF3C

Cl

SO2NH2

N

28

29


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Received: February 20, 2007 Revised: April 13, 2007 Accepted: April 15, 2007

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Development of Non-Nucleoside Reverse Transcriptase Inhibitors for Anti-HIV Therapy