Editing Reverse-transcriptase inhibitor (section)

==Mechanisms of resistance to reverse transcriptase inhibitors==
While NRTIs and NNRTIs alike are effective at terminating DNA synthesis and HIV replication, HIV can and eventually does develop mechanisms that confer the virus resistance to the drugs. HIV-1 RT does not have proof-reading activity. This, combined with selective pressure from the drug, leads to mutations in reverse transcriptase that make the virus less susceptible to NRTIs and NNRTIs.
Aspartate residues 110, 185, and 186 in the reverse transcriptase polymerase domain are important in the binding and incorporation of nucleotides. The side chains of residues K65, R72, and Q151 interact with the next incoming nucleotide. Also important is L74, which interacts with the template strand to position it for base pairing with the nucleotide. Mutation of these key amino acids results in reduced incorporation of the analogs.

===NRTI resistance===
There are two major mechanisms of NRTI resistance. The first being reduced incorporation of the nucleotide analog into DNA over the normal nucleotide. This results from mutations in the N-terminal polymerase domain of the reverse transcriptase that reduce the enzyme's affinity or ability to bind to the drug . A prime example for this mechanism is the M184V mutation that confers resistance to lamivudine (3TC) and emtricitabine (FTC).<ref>{{cite journal |last1 = Hachiya | first1 = A | last2 = Kodama | first2 = EN | last3 = Schuckmann | first3 = MM | last4 = Kirby | first4 = KA | last5 = Michailidis | first5 = E | last6 = Sakagami | first6 = Y | last7 = Oka | first7 = S | last8 = Singh | first8 = K | last9 = Sarafianos | first9 = SG | editor1-last = Ambrose | editor1-first = Zandrea |title=K70Q adds high-level tenofovir resistance to "Q151M complex" HIV reverse transcriptase through the enhanced discrimination mechanism |journal=PLOS ONE |volume=6 |issue=1 |pages=e16242 |year=2011 |pmid=21249155 |pmc=3020970 |doi=10.1371/journal.pone.0016242 | bibcode = 2011PLoSO...616242H | doi-access = free |bibcode-access=free }}</ref><ref>{{cite journal | last1 = Sarafianos | first1 = SG | last2 = Das | first2 = K | last3 = Clark Jr | first3 = AD | last4 = Ding | first4 = J | last5 = Boyer | first5 = PL | last6 = Hughes | first6 = SH | last7 = Arnold | first7 = E | title = Lamivudine (3TC) resistance in HIV-1 reverse transcriptase involves steric hindrance with beta-branched amino acids |bibcode-access=free | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 96 | issue = 18 | pages = 10027–32 | year = 1999 | pmid = 10468556 | pmc = 17836 | doi=10.1073/pnas.96.18.10027| bibcode = 1999PNAS...9610027S | doi-access = free }}</ref> Another well characterized set of mutations is the Q151M complex found in multi-drug resistant HIV which decreases reverse transcriptase's efficiency at incorporating NRTIs, but does not affect natural nucleotide incorporation. The complex includes Q151M mutation along with A62V, V75I, F77L, and F116Y.<ref>{{cite journal | last1 = Shafer | first1 = RW | last2 = Kozal | first2 = MJ | last3 = Winters | first3 = MA | last4 = Iversen | first4 = AK | last5 = Katzenstein | first5 = DA | last6 = Ragni | first6 = MV | last7 = Meyer | first7 = WA III | last8 = Gupta | first8 = P | last9 = Rasheed | first9 = S | last10 = Coombs | first10 = R. | last11 = Katzman | first11 = M. | last12 = Fiscus | first12 = S. | last13 = Merigan | first13 = T. C. | title = Combination therapy with zidovudine and didanosine selects for drug-resistant human immunodeficiency virus type 1 strains with unique patterns of ''pol'' gene mutations | journal = The Journal of Infectious Diseases | volume = 169 | issue = 4 | pages = 722–9 | year = 1994 | pmid = 8133086 | doi = 10.1093/infdis/169.4.722 | display-authors = 8 }}</ref><ref>{{cite journal | last1 = Iversen | first1 = AK | last2 = Shafer | first2 = RW | last3 = Wehrly | first3 = K | last4 = Winters | first4 = MA | last5 = Mullins | first5 = JI | last6 = Chesebro | first6 = B | last7 = Merigan | first7 = TC | title = Multidrug-resistant human immunodeficiency virus type 1 strains resulting from combination antiretroviral therapy | journal = Journal of Virology | volume = 70 | issue = 2 | pages = 1086–90 | year = 1996 | pmid = 8551567 | pmc = 189915 | doi = 10.1128/JVI.70.2.1086-1090.1996 |doi-access=free }}</ref> A virus with Q151M alone is intermediately resistant to zidovudine (AZT), didanosine (ddI), zalcitabine (ddC), stavudine (d4T), and slightly resistant to abacavir (ABC).<ref>{{cite journal | last1 = Maeda | first1 = Y | last2 = Venzon | first2 = DJ | last3 = Mitsuya | first3 = H | title = Altered drug sensitivity, fitness, and evolution of human immunodeficiency virus type 1 with pol gene mutations conferring multi-dideoxynucleoside resistance | journal = The Journal of Infectious Diseases | volume = 177 | issue = 5 | pages = 1207–13 | year = 1998 | pmid = 9593005 | doi = 10.1086/515282 | doi-access =  }}</ref><ref name="Pmid">{{cite journal | last1 = Matsumi | first1 = S | last2 = Kosalaraksa | first2 = P | last3 = Tsang | first3 = H | last4 = Kavlick | first4 = MF | last5 = Harada | first5 = S | last6 = Mitsuya | first6 = H | title = Pathways for the emergence of multi-dideoxynucleoside-resistant HIV-1 variants | journal = AIDS | volume = 17 | issue = 8 | pages = 1127–37 | year = 2003 | pmid = 12819513 | doi =  10.1097/00002030-200305230-00003| s2cid = 21484628 | doi-access = free }}</ref> A virus with Q151M complexed with the other four mutations becomes highly resistant to the above drugs, and is additionally resistant to lamivudine (3TC) and emtricitabine (FTC).<ref name="Pmid" /><ref>{{cite journal | last1 = Gao | first1 = HQ | last2 = Boyer | first2 = PL | last3 = Sarafianos | first3 = SG | last4 = Arnold | first4 = E | last5 = Hughes | first5 = SH | title = The role of steric hindrance in 3TC resistance of human immunodeficiency virus type-1 reverse transcriptase | journal = Journal of Molecular Biology | volume = 300 | issue = 2 | pages = 403–18 | year = 2000 | pmid = 10873473 | doi = 10.1006/jmbi.2000.3823 | url = https://zenodo.org/record/1229892 }}</ref>

The second mechanism is the excision or the hydrolytic removal of the incorporated drug or [[pyrophosphate|pyrophosphorolysis]]. This is a reverse of the polymerase reaction in which the pyrophosphate/PPI released during nucleotide incorporation reacts with the incorporated drug (monophosphate) resulting in the release of the triphosphate drug. This 'unblocks' the DNA chain, allowing it to be extended, and replication to continue.<ref name="Pmid_a">{{cite journal | last1 = Meyer | first1 = PR | last2 = Matsuura | first2 = SE | last3 = Mian | first3 = AM | last4 = So | first4 = AG | last5 = Scott | first5 = WA | title = A mechanism of AZT resistance: an increase in nucleotide-dependent primer unblocking by mutant HIV-1 reverse transcriptase | journal = Molecular Cell | volume = 4 | issue = 1 | pages = 35–43 | year = 1999 | pmid = 10445025  |doi = 10.1016/S1097-2765(00)80185-9 | doi-access = free }}</ref> Excision enhancement mutations, typically M41L, D67N, K70R, L210W, T215Y/F, and K219E/Q, are selected for by thymidine analogs AZT and D4T; and are therefore called thymidine analog mutations (TAMs).<ref name="Pmid_a" /><ref>{{cite journal | last1 = Boyer | first1 = PL | last2 = Sarafianos | first2 = SG | last3 = Arnold | first3 = E | last4 = Hughes | first4 = SH | title = Selective excision of AZTMP by drug-resistant human immunodeficiency virus reverse transcriptase | journal = Journal of Virology | volume = 75 | issue = 10 | pages = 4832–42 | year = 2001 | pmid = 11312355 | pmc = 114238 | doi = 10.1128/JVI.75.10.4832-4842.2001 }}</ref><ref>{{cite journal | last1 = Arion | first1 = D | last2 = Kaushik | first2 = N | last3 = McCormick | first3 = S | last4 = Borkow | first4 = G | last5 = Parniak | first5 = MA | title = Phenotypic mechanism of HIV-1 resistance to 3'-azido-3'-deoxythymidine (AZT): increased polymerization processivity and enhanced sensitivity to pyrophosphate of the mutant viral reverse transcriptase | journal = Biochemistry | volume = 37 | issue = 45 | pages = 15908–17 | year = 1998 | pmid = 9843396 | doi = 10.1021/bi981200e }}</ref> Other mutations including insertions and deletions in the background of the above mutations also confer resistance via enhanced excision.<ref name="Pmid" />

===NNRTI resistance===
NNRTIs do not bind to the active site of the polymerase but in a less conserved pocket near the active site in the p66 subdomain. Their binding results in a conformational change in the reverse transcriptase that distorts the positioning of the residues that bind DNA, inhibiting polymerization.<ref>{{cite journal | last1 = De Clercq | first1 = E | title = The role of non-nucleoside reverse transcriptase inhibitors (NNRTIs) in the therapy of HIV-1 infection | journal = Antiviral Research | volume = 38 | issue = 3 | pages = 153–79 | year = 1998 | pmid = 9754886 | doi = 10.1016/S0166-3542(98)00025-4 }}</ref> Mutations in response to NNRTIs decrease the binding of the drug to this pocket. Treatment with a regimen including efavirenz (EFV) and nevirapine (NVP) typically results in mutations L100I, Y181C/I, K103N, V106A/M, V108I, Y188C/H/L and G190A/S.<ref>{{cite journal | last1 = Johnson | first1 = VA | last2 = Brun-Vezinet | first2 = F | last3 = Clotet | first3 = B | last4 = Gunthard | first4 = HF | last5 = Kuritzkes | first5 = DR|author5-link=Daniel Kuritzkes | last6 = Pillay | first6 = D | last7 = Schapiro | first7 = JM | last8 = Richman | first8 = DD | title = Update of the drug resistance mutations in HIV-1: December 2009 | journal = Topics in HIV Medicine | volume = 17 | issue = 5 | pages = 138–45 | year = 2009 | pmid = 20068260 }}</ref>
There are three main mechanisms of NNRTI resistance. In the first NRTI mutations disrupt specific contacts between the inhibitor and the NNRTI binding pocket. An example of this is K103N and K101E which sit at the entrance of the pocket,<ref>{{cite journal | doi = 10.1016/j.jmb.2006.08.097 | last1 = Das | first1 =   Kalyan| year = 2007 | last2 = Sarafianos | first2 = SG | last3 = Clark Jr | first3 = AD | last4 = Boyer | first4 = PL | last5 = Hughes | first5 = SH | last6 = Arnold | first6 = E | title = Crystal structures of clinically relevant Lys103Asn/Tyr181Cys double mutant HIV-1 reverse transcriptase in complexes with ATP and non-nucleoside inhibitor HBY 097 | journal = J Mol Biol | volume = 365 | issue = 1| pages = 77–89 | pmid = 17056061 }}</ref><ref>{{cite journal | last1 = Hsiou | first1 = Y | last2 = Ding | first2 = J | last3 = Das | first3 = K | last4 = Clark Jr | first4 = AD | last5 = Boyer | first5 = PL | last6 = Lewi | first6 = P | last7 = Janssen | first7 = PA | last8 = Kleim | first8 = JP | last9 = Rösner | first9 = M | last10 = Hughes | first10 = Stephen H | last11 = Arnold | first11 = Edward | title = The Lys103Asn mutation of HIV-1 RT: a novel mechanism of drug resistance | journal = Journal of Molecular Biology | volume = 309 | issue = 2 | pages = 437–45 | year = 2001 | pmid = 11371163 | doi = 10.1006/jmbi.2001.4648 | s2cid = 3109889 | display-authors = 8 }}</ref> blocking the entrance/binding of the drug. A second mechanism is the disruption of important interactions on the inside of the pocket. For example, Y181C and Y188L result in the loss of important aromatic rings involved in NNRTI binding.<ref>{{cite journal | last1 = Ren | first1 = J | last2 = Nichols | first2 = C | last3 = Bird | first3 = L | last4 = Chamberlain | first4 = P | last5 = Weaver | first5 = K | last6 = Short | first6 = S | last7 = Stuart | first7 = DI | last8 = Stammers | first8 = DK | title = Structural mechanisms of drug resistance for mutations at codons 181 and 188 in HIV-1 reverse transcriptase and the improved resilience of second generation non-nucleoside inhibitors | journal = Journal of Molecular Biology | volume = 312 | issue = 4 | pages = 795–805 | year = 2001 | pmid = 11575933 | doi = 10.1006/jmbi.2001.4988 }}</ref><ref>{{cite journal | last1 = Das | first1 = K | last2 = Ding | first2 = J | last3 = Hsiou | first3 = Y | last4 = Clark Jr | first4 = AD | last5 = Moereels | first5 = H | last6 = Koymans | first6 = L | last7 = Andries | first7 = K | last8 = Pauwels | first8 = R | last9 = Janssen | first9 = PA | last10 = Boyer | first10 = Paul L. | last11 = Clark | first11 = Patrick | last12 = Smith | first12 = Richard H. | last13 = Kroeger Smith | first13 = Marilyn B. | last14 = Michejda | first14 = Christopher J. | last15 = Hughes | first15 = Stephen H. | last16 = Arnold | first16 = Edward | title = Crystal structures of 8-Cl and 9-Cl TIBO complexed with wild-type HIV-1 RT and 8-Cl TIBO complexed with the Tyr181Cys HIV-1 RT drug-resistant mutant | journal = Journal of Molecular Biology | volume = 264 | issue = 5 | pages = 1085–100 | year = 1996 | pmid = 9000632 | doi=10.1006/jmbi.1996.0698| display-authors = 8 }}</ref> The third type of mutations result in changes in the overall conformation or the size of the NNRTI binding pocket. An example is G190E, which creates a steric bulk in the pocket, leaving little or no room for an NNRTI to tightly bind.<ref>{{cite journal | last1 = Hsiou | first1 = Y | last2 = Das | first2 = K | last3 = Ding | first3 = J | last4 = Clark Jr | first4 = AD | last5 = Kleim | first5 = JP | last6 = Rösner | first6 = M | last7 = Winkler | first7 = I | last8 = Riess | first8 = G | last9 = Hughes | first9 = SH | last10 = Arnold | first10 = Edward | title = Structures of Tyr188Leu mutant and wild-type HIV-1 reverse transcriptase complexed with the non-nucleoside inhibitor HBY 097: inhibitor flexibility is a useful design feature for reducing drug resistance | journal = Journal of Molecular Biology | volume = 284 | issue = 2 | pages = 313–23 | year = 1998 | pmid = 9813120 | doi = 10.1006/jmbi.1998.2171 | display-authors = 8 }}</ref><ref>{{cite journal | last1 = Ren | first1 = J | last2 = Esnouf | first2 = R | last3 = Garman | first3 = E | last4 = Somers | first4 = D | last5 = Ross | first5 = C | last6 = Kirby | first6 = I | last7 = Keeling | first7 = J | last8 = Darby | first8 = G | last9 = Jones | first9 = Y | last10 = Stuart | first10 = David | last11 = Stammers | first11 = David | title = High resolution structures of HIV-1 RT from four RT-inhibitor complexes | journal = Nature Structural Biology | volume = 2 | issue = 4 | pages = 293–302 | year = 1995 | pmid = 7540934 | doi = 10.1038/nsb0495-293 | s2cid = 34618424 | display-authors = 8 }}</ref>