Tenofovir has been proposed as a drug of choice in antiviral drug strategies directed against the global pandemic , as a result of its favourable resistance profile, long intracellular half-life, and marginal adverse effects [17–20]. Current guidelines recommend tenofovir/emtricitavudine (Truvada) or zidovudine/lamivudine (Combivir) with efavirenz as two first-line treatments of choice for antiretroviral-naive patients . Clinical trials to evaluate the efficiency of tenofovir in pre-exposure prophylaxis have been initiated in Africa, Asia, and the United States . Among the reasons for choosing tenofovir over such drugs as lamivudine has been the reduced risk of developing resistance to tenofovir via the K65R mutation.
We have now demonstrated that tenofovir can rapidly select K65R in subtype C viruses in cell culture. The appearance of K65R with tenofovir is as rapid as that of M184V with lamivudine. These findings indicate that tenofovir-based regimens in subtype C infections will need to be carefully monitored. A recent report suggested that K65R is less prevalent in patients harbouring subtype A than in subtype B and C infections . On the basis of our findings, the reduced prevalence of K65R in subtype A may partly be explained by a higher rate of selection in subtype C infections.
The mechanisms involved in the more rapid selection of K65R among subtype C viruses may relate to the sequences that naturally encode a wild-type KKK motif at codons 64, 65 and 66. Only a single point mutation is required for the emergence of K65R in each of the viral subtypes evaluated. Conceivably, the nucleotide environment of the position 65 codon plays a role in governing the frequency of K65R substitutions, allowing for the more rapid amplification of K65R viruses under conditions of drug pressure. It does not appear that K65R subtype C variants are less impacted in terms of replication capacity, compared with wild-type, than subtype B viruses containing this same substitution .
There are potential limitations in extrapolating in-vitro selection data to the in-vivo situation. Site-directed mutagenesis studies in which the AAA codon 65 of a subtype B clone is replaced by AAG and in which the AAG at codon 65 in subtype C viruses is replaced by AAA are ongoing. Previous work showing that a V106M substitution in subtype C viruses is selected by efavirenz provides proof of concept that in-vitro findings may be reflected in the clinical setting . Several recent studies have shown non-responsiveness to tenofovir/lamivudine/abacavir regimens, leading to the rapid appearance of K65R in 48–68% of patients [24,25]. In contrast, the 934 study showed clinical benefit of a tenofovir/emtricitabine/efavirenz regimen with no appearance of K65R . Such studies demonstrated that drug potency, low genetic barrier to tenofovir resistance, and physiological factors may all contribute to differential responsiveness to tenofovir . Our study highlights the need for careful evaluation of tenofovir-based regimens in subtype C infections.
Although the K65R mutation remains relatively uncommon, an increase in its prevalence in western countries has been observed over the past 4 years, as a result of the increased clinical use of tenofovir [26,28–30]. The K65R mutation may also be associated with the Q151M resistance pathway . Our group has shown with others that three out of eight treated patients in Botswana developed K65R with didanosine-containing regimens . Of note is the fact that the the four subtype C isolates selected with didanosine in this study did not develop K65R, although M184V and L74V were observed in two selections.
It is still unknown whether K65R will become increasingly prevalent in subtype C infections after the usage of tenofovir. Our findings indicate the need for vigilance in adapting antiretroviral regimens and prophylactic schedules for use in developing countries, and that subtype differences in regard to the development of drug resistance need to be carefully monitored whenever antiretroviral drugs are introduced.
1. Spira S, Wainberg MA, Loemba H, Turner D, Brenner BG. The impact of clade diversity on HIV-1 virulence, antiretroviral drug sensitivity and drug resistance. J Antimicrob Chemother 2003; 51:229–240.
2. Essex M, Mboup S. Regional variations in the African epidemics. In: Essex M, Mboup S, Kanki PJ, Marlink RG, Tlou SD, editors. AIDS in Africa. New York: Kluwer/Plenum Publishers; 2002.
3. Kantor R, Katzenstein DA, Efron B, Carvalho AP, Wynhoven B, Cane P, et al
. Impact of HIV-1 subtype and antiretroviral therapy on protease and reverse transcriptase genotype: results of a global collaboration. PLOS Med 2005; 2:325–337.
4. Osmanov S, Pattou C, Walker N, Schwardlander B, Esparza J. WHO–UNAIDS Network for HIV Isolation and Characterization. Estimated global distribution and regional spread of HIV-1 genetic subtypes in the year 2000. J Acquir Immune Defic Syndr 2002; 29:184–190.
5. Brenner B, Turner D, Oliveira M, Moisi D, Detorio M, Carobene M, et al
. A V106M mutation in HIV-1 clade C viruses exposed to efavirenz confers cross-resistance to non-nucleoside reverse transcriptase inhibitors. AIDS 2003; 17:F1–F5.
6. Abecasis AB, Deforche K, Snoeck J, Bacheler L, McKenna P, Carvalho AP, et al
. Protease mutation M89I/V is linked to therapy failure in patients infected with the HIV-1 non-B subtypes C. F or G AIDS 2005; 19:1799–1806.
7. Eshleman S, Guay LA, Wang J, Mwatha A, Brown ER, Musoke P, et al
. Distinct Patterns of emergence and fading of K103N and Y181C in women with subtype A vs. D after single-dose nevirapine: HIVNET 012. J Acquir Immune Defic Syndr 2005; 40:24–29.
8. Gupta RK, Chrystie IL, O'Shea S, Mullen JE, Kulasegaram R, Tong CYW. K65R and Y181C are less prevalent in HAART-experienced HIV-1 subtype A patients. AIDS 2005; 19:1916–1919.
9. Grossman Z, Istomin V, Averbach D, Lorber M, Risenberg K, Levi I, et al
. Genetic variation at NNRTI resistance-associated positions in patients infected with HIV-1 subtype C. AIDS 2004; 18:909–915.
10. Grossman Z, Paxinos EE, Averback D, Maayan S, Parkin NT, Engelhard D, et al
. Mutation D30N is not preferentially selected by human immunodeficiency virus type 1 subtype C in the development of resistance to nelfinavir. Antimicrob Agents Chemother 2004; 48:2159–2165.
11. Turner D, Brenner B, Moisi D, Detorio M, Cesaire R, Kurimuri T, et al
. Nucleotide and amino acid polymorphisms at drug resistance sites in non-B subtype HIV-1 variants. Antimicrob Agents Chemother 2004; 48:2993–2998.
12. Wainberg MA, Miller MD, Quan Y, Salomon H, Mulato AS, Lamy PD, et al
. In vitro selection and characterization of HIV-1 with reduced susceptibility to PMPA. Antiviral Ther 1999; 4:87–94.
13. Loemba H, Brenner B, Parniak MA, Ma'ayan S, Spira B, Moisi D, et al
. Polymorphisms of cytotoxic T-lymphocyte (CTL) and T-helper epitopes within reverse transcriptase (RT) of HIV-1 subtype C from Ethiopia and Botswana following selection of antiretroviral drug resistance. Antiviral Res 2002; 56:129–142.
14. Loemba H, Brenner B, Parniak MA, Ma'ayan S, Spira B, Moisi D, et al
. Genetic divergence of HIV-1 Ethiopian clade C reverse transcriptase (RT) and rapid development of resistance against non-nucleoside inhibitors of RT. Antimicrob Agents Chemother 2002; 46:2087–2094.
15. Salomon H, Belmonte A, Nguyen K, Gu Z, Gelfand M, Wainberg MA. Comparison of cord blood and peripheral blood mononuclear cells as targets for viral isolation and drug sensitivity studies involving human immunodeficiency virus type 1. J Clin Microbiol 1994; 32:2000–2002.
16. Wainberg MA. Generic HIV drugs-enlightened policy for global health. N Engl J Med 2005; 352:747–750.
17. De Clercq E, Holy A. Acyclic nucleoside phosphonates: a key class of antiviral drugs. Nat Rev 2005; 4:928–940.
18. Gallant JE, Deresinski S. Tenofovir disproxyl fumarate. Clin Infect Dis 2003; 37:849–859.
19. Gallant JE, Staszewski S, Pozniak AL, DeJesus E, Sulieman JM, Miller MD, et al
. Efficacy and safety of tenofovir DF vs stavudine in combination therapy in antiretroviral-naive patients. A 3-year randomized trial. JAMA 2004; 292:191–201.
20. Barditch-Crovo P, Deeks SG, Collier A, Safrin S, Coakley DF, Miller M, et al
. Phase i/ii trial of the pharmacokinetics, safety, and antiretroviral activity of tenofovir disoproxil fumarate in human immunodeficiency virus-infected adults. Antimicrob Agents Chemother 2001; 45:2733–2739.
21. Guidelines for the use of antiretroviral agents in HIV-1-infected adults and adolescents.
Available at: http://AIDSinfo.nih.gov
. Accessed: 6 October 2005.
22. Grant RM, Buchbinder S, Cates W Jr, Clarke E, Coates T, Cohen MS, et al
. Promote HIV chemoprophylaxis research, don't prevent it. Science 2005; 309:2170–2171.
23. White KL, Margot NA, Wrin T, Petropoulos CJ, Miller MD, Naeger LK. Molecular mechanisms of resistance to human immunodeficiency virus type 1 with reverse transcriptase mutations K65R and K65R+M184V and their effects on enzyme function and viral replication capacity. Antimicrob Agents Chemother 2002; 46:3437–3446.
24. Gallant JE, Rodriguez AE, Weinberg WG, Young B, Berger DS, Lim ML, et al
. Early virologic nonresponse to tenofovir, abacavir, and lamivudine in HIV-infected antiretroviral-naive subjects. J Infect Dis 2005; 192:1921–1930.
25. Delauney C, Brun-Vezinet F, Landman R, Collin G, Peytavin G, Trylesinski A, et al
. Comparative selection of the K65R and M184V/I mutations in human immunodeficiency virus type 1-infected patients enrolled in a trial of first-line triple-nucleoside analog therapy (Tonus IMEA 021). J Virol 2005; 79:9572–9578.
26. Gallant JE, DeJesus E, Arribas JR, Pozniak AL, Gazzard B, Campo RE, et al
. Tenofovir DF, emtricitabine, and efavirenz vs. zidovudine, lamivudine, and efavirenz for HIV. N Engl J Med 2006; 354:251–260.
27. Kuritkes DR. Less than the sum of its parts: failure of a tenofovir-abacavir-lamivudine triple-nucleoside regimen. J Infect Dis 2005; 192:1867–1868.
28. Winston A, Pozniak A, Mandalia S, Gazzard B, Pillay D, Nelson M. Which nucleoside and nucleotide backbone combinations select for the K65R mutation in HIV-1 reverse transcriptase. AIDS 2004; 18:949–951.
29. Valer L, Martin-Carbonero L, de Mendoza C, Corral A, Soriano V. Predictors of selection of K65R: tenofovir use and lack of thymidine analogue mutations. AIDS 2004; 18:2094–2096.
30. Gianotti N, Seminari E, Fusetti G, Salpietro S, Boeri E, Galli A, et al
. Impact of a treatment including tenofovir plus didanosine on the selection of the 65R mutation in highly drug-experienced HIV-infected patients. AIDS 2004; 18:2205–2208.
31. Doualla-Bell F, Gaseitsiwe S, Ndung'u T, Modukanele M, Peter T, Novitsky V, et al
. Mutations and polymorphisms associated with antiretroviral drugs in HIV-1C-infected African patients. Antivir Chem Chemother 2004; 15:189–200.