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  11th Intl. HIV Drug Resistance Workshop
July 2-5, 2002
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Summary Report on the Resistance Workshop
Written by David Margolis, MD, University of Texas Southwestern Medical Center
  Resistance to New Antiretroviral Agents
Merck Integrase Inhibitors splash onto the scene
The session opened with impressive back-to-back presentations by Daria Hazuda and Stephen Young of the Merck HIV Integrase Inhibitor Discovery Team. Ending 2 years of troubling silence since this group first described the antiretroviral activity of diketobutanoic acids (DKAs) that specifically and potently inhibit the critical strand transfer step of the integration of the HIV provirus into the human genome, Hazuda and Young drew back the curtain to reveal oral integrase inhibitors ready for Phase I human study.
The integrase enzyme of HIV mediates the splicing of the HIV genome into the host cell DNA and is required for replication. Great excitement was generated when Hazuda's group first described the specific mechanism by which DKAs inhibited integration in 2000, but until these presentations there was a growing concern about the lack of reported progress towards a testable drug.
Hazuda reported that as the original DKAs were found to have unfavorable pharmacokinetic properties, a related class of compounds, 1,6 napthyridine-7-carboxamides, had been developed. These drugs were found to be orally bioavailable and inhibit 50% of HIV replication in culture at a concentration of 0.25 M. This respectable IC50 (inhibitory concentration 50%) was found in the presence of 50% human serum.
Hazuda then leapfrogged the usual reporting of drug efficacy and toxicity studies in culture and animal systems, and revealed that the pilot compound L,870,812 was effective in a primate model of HIV infection. Rhesus macaques were infected with a chimeric SIV-HIV (SHIV 89.6P). This viral strain in this animal induces rapid CD4 depletion within weeks. Monkeys were treated with 810 alone shortly after infection. Plasma viral loads declined from 1 to 3 logs, and up to 4 logs in some animals when declines were measured with a more sensitive assay. CD4 counts were preserved during more than 10 weeks of treatment. Although the true potency of this drug might be overrepresented by its testing in the setting of acute infection, this model allowed a rapid proof of concept of antiviral activity.
Young then expanded on the properties of L,870,810, the drug selected for first human testing (Merck is cleverly developing two top candidates in parallel, so that there is a back-up ready if one gets tripped up). He reported that in cell culture assays, 810 induced declines in integrated HIV DNA in concert with increases of dead-end, unintegrated circularized HIV genomes (known as 2-LTR circles). Exposure to 810 induced mutations in the catalytic core of HIV integrase, resulting in 1.6-fold to 9-fold resistance. The IC95 of 810 was ca. 15-100 nM against a variety of HIV strains in lymphocytes and macrophages, and the IC50 in culture was only 4 nM against a virus clone resistant to all currently licensed classes of antiretrovirals -- meaning the drug should not be cross-resistant with currently available classes of drugs. Pharmacokinetic studies show that 810 is metabolized by glucuronidation, not the hepatic p450 system as are protease inhibitors. The drug's half-life is 2.3 hours, allowing twice a day dosing. It was said that 810 has "moved into Phase I testing in healthy volunteers."
FLT (MIV-310 or alovudine): a drug unearthed for a second coming?
3'-deoxy-3'-fluorothymidine, a drug briefly tested in the 1980's, has been dusted off and retested in a cohort of patients with multiple nucleoside analog mutations. In earlier days, when used as monotherapy this drug was not better than ZDV, caused low platelet counts, and so fell by the wayside. As reported by Vincent Calvez, due to in vitro activity against multinucleoside resistant strains, MIV-310 7.5 mg qd was added to the therapy of 15 patients with >2 nucleoside analog mutations, and median 3.93 log copies HIV RNA. A decline of 1.13 log copies of HIV RNA was seen at 4 weeks. A greater decline (median 1.88 log) was seen in patients not receiving stavudine, raising the possibility of an as-yet unknown interaction with or competition between D4T and MIV-310. Although some in the audience reminded the group of their unhappy experience with the drug years ago, others felt that it was time to give a drug with activity against MDR virus a second chance. Perhaps in the context of combination therapy and better management of toxicities, FLT can make a comeback.
TMC 114: a tougher non-nuke to come?
The heartening procession of new drug candidates continued with discussion of Tibotec's protease inhibitor TMC 114. A potent PI, with a 50% effective concentration of 10 nM, de Bethune presented her group's difficulty in selecting TMC 114- resistant HIV in culture. This was contrasted with the ease in which resistance was selected in her system to current PIs. We look forward to human testing of this drug.
Laboratory studies of the Tibotec non-nucleoside RT inhibitor, TMC 125 were also presented. These studies showed preserved activity of TMC 125 despite the presence of the NNRTI class-killer mutation K103N. In the presence of modest concentrations of TMC, higher-level resistance viruses did not emerge in culture if the infecting strain contained only K103N. If the infecting strain contained the double mutant K103N/Y181C, TMC 125 resistance developed in a manner comparable to current NNRTIs. Although this data did suggest that TMC 125 posed a higher genetic barrier to NNRTI resistance, many patients with NNRTI exposure do carry double mutations within the NNRTI class. So while the barrier to resistance may be higher, the durable utility of TMC 125 in NNRTI-experienced populations is still to be tested. Some clinical evidence to support this claim is to be presented at Barcelona. Further clinical development of this promising drug is awaited.
BMS-806: the potential to hit the HIV envelope pocket that grabs CD4
Ping-Fan Lin presented the surprising initial success of a small, orally bioavailable molecule, BMS-806, which binds the HIV envelope glycoprotein gp120. BMS-806 binds gp120 within the pocket that gp120 uses to bind CD4. Binding resulted in inhibition of lab and clinical isolates at EC50s of 2.5-61 nM. Although the outgrowth of drug-resistant viruses with mutations in the gp120 region bound by the drug were reported, the possibility of another tool to inhibit HIV entry by yet another mechanism was encouraging.
Although small molecules which bind to the gp120 envelope protein of HIV are likely to be susceptible to escape by viral variation and mutation, if such drugs are developed they may present several problems for HIV. First, as such drugs may be targeted to conserved and functionally important sites of the envelope protein such as the CD4 binding pocket, viral variation at such a region may come at a considerable cost to the virus. Such mutant viruses may not be able to replicate as well, termed a loss of "fitness." There is already evidence that some mutations that allow resistance to current antiretrovirals come at a fitness cost to the virus, and that a loss of viral fitness might be associated with a clinical benefit.
Perhaps more significantly, as fusion inhibitors, anti-gp120 drugs, and chemokine receptor antagonists are developed, therapeutic regimens may emerge that are capable of simultaneously constraining the viral fusion/entry process from multiple directions. Viral envelope (gp120 and gp41), which mediates CD4 and chemokine receptor interaction and viral binding/fusion, is also the primary target of the antiviral immune response. This is new and important, as the current targets of drug therapy, the RT and protease proteins, are poorly immunogenic and are not primary targets of the immune response. It is conceivable that clever use of these new classes of drugs may force the viral envelope mutation along pathways that may be better controlled by the immune response. Thought of another way, chemotherapies that target envelope are assisted by another powerful "drug", the immune system.
Tipranavir: still hoping for a super-PI
Resistance information was reported on the non-peptidic protease inhibitor tipranavir (TPV) from a phase II open-label study. Due to study design it is thus far difficult to discern the activity of TPV. 41 patients with exposure to multiple PIs, but who were NNRTI naive, were treated for up to 48 weeks with one of two doses of TPV boosted by ritonavir, with efavirenz and one new NRTI. During the trial the TPV capsule formulation was changed in all patients. All but 2 subjects had between 6 and 20 PI mutations at baseline. During the 48 weeks of the study, 5 of the 41 patients had an increase in phenotypic (Virco) resistance to TPV. In these subjects, with an average of 16 PI mutations at baseline, most developed novel mutations at V82T and at L33 (to V,F, or I) in association with decreased sensitivity to TPV. Studies are now underway to determine the optimal dose of TPV, to allow definitive testing of this long-awaited drug.
The fusion inhibitors T-20 and T-1249
Several presentations were made by the Trimeris group throughout the meeting, expanding our information about the novel fusion inhibitor now in Phase III study, T-20, and the potent related molecule T-1249. Resistance can develop to either T-20 or T-1249 via mutations in the gp41 ectodomain region AA 35-46, the region that is targeted by T-20 and T-1249. Some mutations that exert effect on T-20 do not induce resistance to T-1249, suggesting that resistance is similar but not identical between the 2 drugs. Loss of T-20 mutations after a mean of 4 months (range 2.4-8.7 months) in patients who stopped T-20 after reaching the end of a protocol gave rise to the suggestion that T-20 resistance mutations come at a (fitness) cost to the virus' ability to replicate.
Taken as a group, in three Phase II T-20 studies, 42/168 subjects met protocol-defined failure criteria. Of these, 40 could be genotyped, and 31 (78%) carried viruses with mutations in the 35-46 region. Most of these exhibited a more than 10-fold decrease in susceptibility to T-20.
Double mutations within this critical gp41 ectodomain region found in patients induced more resistance in vitro than single mutations. In 19 viral isolates from patients that were treated with T-20, sensitivity to T-20 decreased 20-fold, but only 3-fold to T-1249.
In general, at baseline the EC50 of most natural isolates to T-20 is well below 0.1 g/ml, but in some isolates it is as high as 0.5 g/ml. Patient virus is more sensitive to T-1249 at baseline, as the EC50 of most isolates is less than 10 ng/ml, and up to 50 ng/ml.
Some results of a T-1249 trial in patients was also first reported. Across a range of doses, average decline in plasma viral RNA was dose proportional. Mutations in resistant viruses isolated were found in the AA 36-75 region, as expected. Preliminarily, 2 mutations appear needed to induce high-level resistance to T-1249.
Zinc finger inhibitors: another potential new drug class
An update on the development of zinc finger inhibitors was presented by Bill Rice. Rice was among the original group of investigators at the NCI who first proposed therapeutics directed at the zinc finger motif of HIV in the 1990's. Zinc fingers are a molecular structural motif in which loops of amino acid are held together by the electrostatic attraction of a single zinc ion. This creates a protruding "finger" structure. HIV uses zinc fingers to hold together its nucleocapsid protein, which performs several irreplaceable steps in the viral lifecycle. Nucleocapsid zinc finger inhibitors were developed at the NCI which had selective effect on the zinc fingers of retroviruses, but not human zinc fingers. These molecules had poor pharmacokinetic properties, however, and are being pursued at the NCI for non-antiretroviral applications such as viral inactivation for antigen preparation.
Rice has since left the NCI, and started Achillion Pharmaceuticals to develop therapeutic zinc finger inhibitors. He presented data showing antiviral activity of a chemically distinct class of inhibitors reported (but not shown) to have improved pharmacokinetic properties. The development of these agents into effective drugs would be an important advance.
3 July, Day One: Mechanisms of HIV Drug Resistance
Cross-resistance between amprenavir (APV) and lopinavir (LPV)
When APV-resistant virus initially emerges, it is usually not cross-resistant to other PIs. The possibility that LPV induced some resistance to APV, and that the reverse was true was first suggested in data presented at the 10th meeting in Scottsdale. Neil Parkin and colleagues from ViroLogic examined more than 1400 samples that had been tested for genotype and phenotype and found to have at least one PI mutation or reduced susceptibility to one PI. When examining the 13% of these viruses that were classified as sensitive to LPV by genotype but resistant by phenotype, they found that some discordant viruses carried mutations also associated with APV resistance. APV mutations alone did not confer resistance to LPV, as LPV resistance still required 8 or more mutations. However, Parkin concluded that if resistance is present to LPV or APV there might be cross-resistance between APV and LPV unreported by current genotype algorithms. Clinical outcomes data will be required to confirm the relevance of this observation.
Nelfinavir resistance patterns in HIV subtype C
HIV subtype B but not C is predominant in the U.S. or Europe, but C may be found in immigrant populations. However, knowledge of whether viral subtype influences mutational pathways under drug pressure is of growing importance as therapies are increasingly used around the world. Grossman and colleagues from Israel and ViroLogic found that when treated with nelfinavir-containing regimens, only 4.5% of patients with subtype C virus who developed NFV resistance developed the D30N mutation. This mutation has been associated with preserved sensitivity to other PIs. In comparison, approximately 50% of patients in the same clinics with subtype B virus who failed NFV developed D30N, a proportion similar to that seen in subtype B patients in the US.
Nucleoside (thymidine) analog mutations (NAMs or TAMs) occur with equal rates on AZT-based or D4T-based therapy
Johnson, Kuritzkes and ACTG colleagues analyzed genotypes from previous ACTG studies in which AZT/3TC, D4T/3TC, or D4T was given as initial therapy. The emergence of the M41L, D67N, K70R, L210W, T215Y/F, and K219Q mutations was examined. Overall, the rate of detection of these mutations was relatively similar over 72 weeks of study, regardless of whether subjects received AZT or D4T. A trend towards a greater risk of developing 2 of these mutations was seen in subjects receiving AZT/3TC. This study joins several presented over the last 2 years suggesting that NAMs or TAMs can be selected by either AZT or D4T.
Drug resistance and substrate recognition in HIV-1 protease: how does the virus do it?
In an elegant study from the Schiffer group at UMass, the interaction of the crystal structure model of HIV protease with the nine target sites within HIV proteins that protease must cleave was analyzed. The interaction of target cleavage sites with the inactive D25N protease variant or the variant that also encoded the primary resistance mutation V82A. This modeling exercise showed that most of the HIV protein target cleavage sites adopt an asymmetric shape to allow interaction with the mutant protease and do not interact with the V82A mutant residue, while drugs to which the protease is resistant maintain a symmetric shape and do interact with the V82A mutant residue. This finding suggests that new designs of structurally flexible protease inhibitors may allow increased activity against drug-resistant protease.
Sugiura and colleagues from Tokyo and Kawasaki presented an elegant study which further enhanced the understanding of protease inhibitor resistance. In addition to the known mutations within the protease gene, and the mutations at sites within the gag gene cleaved by protease, Sugiura defined other gag mutations outside of the cleavage sites which were required to allow the virus to function and replicate efficiently in the presence of PI resistance mutations. It is likely that when the shape of the protease enzyme changes due to drug resistant mutations, random evolutionary changes in the gag protein are also selected that allow it to fit into the mutant protease.
4 July, Day Two: HIV Pathogenesis, Fitness, and Resistance
On the meeting's second day, a variety of presentations dealt with the complex interplay of HIV's ability to replicate in the presence of antiretrovirals, the cost exacted on the virus by drug resistance, and the impact of drug-resistant virus on the immune system.
Grant of UCSF studied 130 therapy-naive subjects diagnosed with recent infection between 1996 and 2001. They compared the ability of recombinants derived from patient viral RNA to replicate in comparison to a lab strain recombinant. In this surrogate "replication capacity" assay, developed by ViroLogic, growth is not restrained by drugs but compared to the lab standard virus.
Interestingly, higher CD4 counts were found in patients with genotypic evidence of drug resistance (ie acquired drug resistance) or reduced replication capacity. These observations do not prove cause and effect, and could be surrogates for other factors, such as patient demographics. Grant also warned that the benefit of an initially higher CD4 count might be counterbalanced by a poorer response to therapy. Related to this finding, Deenan Pillay (Birmingham, UK) reported no difference in the time to CD4 >350/l or to initiation of therapy in patients infected with drug-resistant viruses than those infected with wild-type HIV. However, it is clear that drug-resistance mutations affect HIV replication capacity in several laboratory systems. Whether measurements of this phenomenon can be translated into a clinically useful tool remains to be seen.
Havlir reported studies of a select group of 8 patients who added ABC to indinavir/efavirenz therapy after 5 years of suppression (< 50 copies/ml) on just those 2 agents. Using a boosted Roche assay (L.O.D 2.5 copies/ml) she observed a drop in "residual viremia" from baselines of 2.7 to 23 copies/ml. The mean VL decline from this low level was 0.6 log copies/ml. This decline in low-level viremia suggested that more potent drug therapy can inhibit ongoing, low-level viral replication in productively infected cells, reactivation viremia emanating for the latent, resting cells reservoir, or both.
Deeks reported findings from his ongoing study of subjects on stable HAART with plasma HIV RNA < 100 copies/ml. Patients were assessed in three groups: a) 77 well-controlled patients with <50 copies/ml; b) 14 "blippers" with transient viremic episodes; and c) 23 subjects with persistent viremia between 50 and 1000 copies/ml. Deeks found CD4 andCD8 cell activation increased in group a, but comparable in groups b and c. The total HIV-specific Elispot peptide response was greater in group a and b than c. As levels of drug resistance increased, replication capacity as per the above assay decreased. Deeks proposed that some of these patients were able to remain stable both due to repriming of the immune response by viremia, and by poor replication of drug-resistant virus.
The ViroLogic phenotype test now includes on its report a value for "replication capacity (RC)." It is still unclear how to use this information, which is derived from a modification of the ViroLogic assay, and reflects replication of the viral reconstructs derived from patient RNA in the company's culture system.
Haubrich (UCSD) examined the RC measured in subjects in the CCTG 575 trial of salvage therapy with or without phenotyping. He found that RC in this cohort was increased when VL was higher, and decreased when CD4 counts increased on therapy. Although the correlation of RC with CD4 increases was not tight, it was the best predictor in a multivariate model. RC also generally declined with resistance to 1-2 PIs, but increased with resistance to 4 or more PIs. While other factors such as the available therapy options might play a more important role, Haubrich suggested that RC might be used in experienced patients to guide the decision to change or continue therapy.
Concerned that treatment interruption might lead to the development of drug-resistant populations not detected in routine assays, Meztner (Erlangen) tested samples from within the Swiss Spanish Interruption (SSIT) study. Using a sensitive PCR assay specifically seeking L90M or M184V mutations that arose during interruptions of 2 weeks or longer, 3 of 25 subjects developed L90M at any time, and 9 of 25 (36%) developed M184V. In total 10 or these 25 subjects (40%) developed minority mutants detected in this assay. While none of these SSIT subjects have yet suffered virologic failure, all were < 50 copies/ml for > 6 months at entry, raising further concerns as to the long-term effect of interruption in suppressed patients.
These findings can be considered together with those of Alan Hance and colleagues at Hopital Bichat Claude Bernard. Using a real-time PCR assay revved up to detect 0.1% minority species (i.e. one mutant needle in 1000 wild-type ones), Hance found that the L90M could be detected as a minority population (of varying size) for 2 to 34 months before appearing in a standard genotype. His findings revealed active and dynamic fluxes of viral populations (and mutations), at levels undetectable by standard assays, during the selective pressure of drug therapy. Subpopulations (of mutations) come and grow, waxing and waning in size. It is important to consider that the presence of a minor subpopulation may not guarantee the development of clinical resistance, and that a clinical endpoint study would be needed to demonstrated the clinical relevance of the detection of minor subpopulations.(edit comment: presumably patients had undetectable HIV viral load)
4 July, Day Two: New Technologies and Interpretations
A wide variety of interpretative systems and comparative evaluations of resistance testing approaches were presented. In a summary overview at the end of the meeting John Mellors spoke for all when he challenged the field to develop improved software analysis tools, and techhniques to evaluation and compare resistance testing systems. One reagent that has been made available for this is a standardized set of resistant viruses to use as a test platform.
DeLuca of Rome's Catholic University presented resistance modeling experiments in which a program operating through the use of "fuzzy logic" was used to develop resistance prediction algorithms. Unlike the fuzzy math that Govenor Bush complained about, fuzzy logic is a computer-based technique that assigns binomial values (0 or 1) to "fuzzy" qualitative predictions (more or less resistant) and repeatedly refines and improves the rules developed using testing sets of known outcomes. DeLuca's program was tested and trained on several data sets of patient genotypes with known virological outcomes at 3 and 6 months. The prediction algorithm derived performed better than those of the Stanford or French ANRS groups. DeLuca reminded the audience that his algorithm had been developed from a relatively small data set, but the technique could be further developed or used to improve existing prediction systems.
Zolopa (Stanford) presented the results of the subtly named GUESS study, in which a group of 12 HIV resistance investigators were challenged to predict phenotype from a genotype. Correct guesses were made about 75% of the time for NNRTIs and 3TC, but only 30-40% of the time for other NRTIs or PIs. The experts tended to agree more with each other than with the phenotype, which could reflect an added value of intuitive clinical judgement not provided by phenotype. The experts tended to agree with one another on recommendations for treatment based on the genotype. Providing further support for the need for expert HIV treaters, panelists assessment of drug activity and recommendations for treatment regimen were generally similar.
A study comparing genotypic (GT), phenotypic (PT), and no resistance (SOC) testing in a cohort of 450 military HIV patients was reported by Wegner. Subjects were randomized, but clinicians accessed and interpreted commercial assays as in general practice. When the Virtual Phenotype genotypic report became available, that was used in the GT arm. There was no difference found in the first endpoint of virologic control at one year of follow-up by Kaplan-Meier analysis. However, in patients with exposure to 4 or more drugs, time to endpoint was significantly longer in the GT and PT arms. In NNRTI-experienced patients, time to failure was longer in the PT arm, with a trend to prolonged suppression that did not achieve significance in the GT arm. This interesting study provided evidence that resistance testing as used in the current therapeutic environment by practicioners may confer benefit to patients with longer histories of drug exposure.
5 July, Day Three: Epidemiology and Clinical Implications of Resistance
DDI and M184V-3TC Resistance
Three studies presented evidence that the184V or 184I mutations, long known to impact DDI activity in culture, did not have a clinically significant effect on DDI efficacy. Winters (Stanford) used data from a study in which patients initially treated with a regimen including 3TC switched to DDI/D4T or 3TC/DDI and other drugs. Among patients with 184V or I, the risk of failure increased 2.67 fold for those not switchin to the DDI regimen. As further evidence that the 184 mutation does not increase the ability of the virus to replicate in the face of DDI, most of those who failed in the DDI arm had lost the 184 mutation.
Eron (UNC) presented an analysis of an old ACTG study who now-outdated design allowed him to directly address this issue. 3TC-experienced patients took either DDI or DDI/HU. There was no significant difference in viral load declines in subjects with 3TC experience, whether or not they had the 184 mutation. Viral load declines were also not significantly different than seen in 3TC-naive subjects in another arm of the study.
Pozniak (Chelsea & Westminster Hospital) found that 176 subjects with 184V who were switched to a DDI-containing regimen had an improved virological outcome when compared to patients switched who did not have 184V.
The footprints of nucleoside resistance at the 215 site
When drug pressure to maintain the 215 Y or F mutation is removed, the virus often back-mutates. The resulting amino acid residue (A, C, D, E, N, V, or S) does not confer resistance but is evidence that the old resistant strain is hiding somewhere. Bolotta (Milan) and Lanier (Glaxo SmithKline) both presented evidence from a small number of patients culled from larger cohorts, that the presence of these revertants (and therefore a hidden archive of resistant viruses) conferred an increased risk of subsequent virological failure. 8 of 12 (67%) of the Italian patients, and 6 of 16 (37.5%) of the subjects from the GSK archives failed subsequent therapy. While the magnitude of this risk is difficult to determine from these small samples, it is clear that the risk is increased and that caution is warranted.
Tenofovir resistance defined
An update was presented of the analysis of tenofovir (TDF) resistance obtained from subject within Gilead TDF studies 902 and 907 (333 subjects). In these placebo-controlled, randomized studies, TDF was added to stable background therapy, allowing the isolated effect of TDF to be ascertained.
Multivariate analysis assessed the impact of individual RT mutations at 41, 67, 70, 210, 215, 219 (the so-called TAMs or NAMs, see below), other RT mutations (39, 43, 44, 67, 69, 75, 118, and 208), HIV RNA, CD4, and current and prior therapy. Two mutational "pathways" were seen: 41-210-215 and 67-70-219. The 41-210-215 pathway was highly associated with TDF, the highest levels being seen when these were presented with 2 other of the above RT mutants.
Multiple mutations in the presence of the 67-70-219 pathway were not associated with loss of TDF response. The K67R mutation was associated with TDF resistance and a loss of response, but this mutation was only observed in 6 subjects. As had been previously suggested, the M184V mutation was associated with a slightly improved TDF response, but only improved by -0.12 log.
Closely reproducing Miller's findings, an independent study by Schmidt and colleagues in Europe found that a core set of TAMs (41 and 215) contributed to TDF resistance. She suggested that 3TC resensitized HIV to TDF, and that the drugs should be used together. She also highlighted the role of K65R and L210W in predicting TDF resistance.
Similarly, Masquelier (Bordeaux) scored patients on the basis of presence or absence of a set of mutations. The previously described TAMs 41L, 67N, 210W, and 215Y or F, as well as 44D, 69 D/N/S, and 118I were counted. A score of less than 3 correlated with a VL decline of 1.3 logs, 3-4 with a VL decline of 0.8 logs, and more than 4 an insignificant decline of 0.4 logs. Score was predictive independent of baseline VL, and the presence or absence of an NNRTI or > 2 PI mutations.
Later in the meeting, Miller presented a clinical correlation of the response rates associated with phenotypic resistance by the ViroLogics and Virco assays. Again taken from 112 phenotypes obtained from some of the 157 subjects with Gilead 907, this used the analytical technique of recursive partitioning to determine phenotypic resistance levels at which TDF response becomes unlikely.
Analysis revealed that at up to 1.4-fold resistance (ViroLogic), there was no reduction in TDF response (70% of subjects had > 0.3 log VL decline and median response at week 24 was -0.77 log copies HIV RNA). Response was reduced between 1.4-fold and 4-fold resistance (23% of subjects had > 0.3 log VL decline and median response at weektt 24 was -0.37 log copies HIV RNA), and was lost above 4-fold resistance (7% of subjects had > 0.3 log VL decline and median response at week 24 was -0.06 log copies HIV RNA). Using the Virco assay, more than 4-fold resistance also correlated with a lack of response. Similar to the recent analysis of this type performed for abacavir resistance, these data give clear guidance to clinicians on the use of genotypic and phenotypic testing with TDF.
Mutations induced by tenofovir (TDF) with or without abacavir (ABC) or 3TC
K65R is a resistance mutation within RT that is selected by both TDF and ABC, but is thus far not seen commonly in clinical practice. To model the use of these drug together in vitro, dose-escalating in vitro passage was performed by Stone and colleagues at Glaxo UK. They found that, as expected, ABC alone selects for mutations at 65, 74, 115, and 184. TDF alone selects for mutations at 62 and 65. However, TDF and ABC together selected for K65R alone, as did TDF and 3TC.
In another presentation, Ait-Khalid and colleagues at GSK US and UK, found that is clinical trials in which ABC or ABC/3TC was the only NRTI given, K65R appeared in 10% and L74V appeared in 40% of 127 subjects failing therapy. However, when ABC was given with zidovudine K65R appeared in 1% and L74V appeared in 2% of 86 subjects failing therapy.
These in vitro and clinical findings serve notice that as novel NRTI combinations are used broadly in the clinic, unexpected or previously uncommon pathways of resistance may be observed. Although the use of potent and simplified therapies is certainly in the best interest of patients, careful attention should be paid to the resistance patterns that emerge.
Transmission of drug-resistant HIV: no bad news yet
A variety of studies on the incidence of drug resistance mutations from cohorts from around the world were presented. The multicenter study led by Little (UCSD) of phenotypic resistance in drug-naive patients found that the incidence of phenotypic resistance to any drug class had decreased from 12.2% to 6.7%. The incidence of resistance to more than one drug class had decreased from 6.5% to 0.7%. At the very least this is not bad news, as while the confidence intervals of these measurements is not large enough to say that the incidence of resistance is decreasing, it seems likely that there has not been a significant increase in resistance over the last year.
Other reports of the incidence of genotypic resistance from the US, Canada, and Europe found an incidence of single-class mutations of 6 to 10%, but for Pillay's study from England which found an incidence of 22%. Samples for these studies were obtained between 1994 and 2001, with the majority being obtained after 1999. When reported, resistance to more than one class was rare (0.2 to 0.7%), although Pillay's cohort again found an incidence of 1.5%.