Higher intracellular concentrations with tenofovir alafenamide (TAF) overcomes K65R and other key NRTI resistance in vitro
Simon Collins, HIV i-Base
Results of a careful study suggest that the new tenofovir prodrug will become an essential goal for global treatment for at least the next decade. 
The study was presented at the International Workshop on HIV and Hepatitis Drug Resistance held in Toronto from 6-8 June 2013. 
Tenofovir alafenamide (TAF) is a prodrug that has greater viral activity compared to tenofovir disoproxil fumerate (TDF) – by approximately an additional 0.5 log copies/mL in treatment naive patients – and which achieves intracellular concentrations of tenofovir diphosphate in PBMCs that are 5-7 fold higher and tenofovir (TFV) plasma concentrations that are 90% lower.
First in vitro results on the resistance profile for TAF suggest that this improved cell targeting results in a resistance profile that overcomes common NRTI resistance including the K65R mutation associated with high level resistance to tenofovir, the MDR Q151M and T69ins mutations and other mutation combinations including thymidine analogue mutations (TAMs). This is based on TAF achieving intracellular 95% inhibitory quotient (IQ95) levels that are five times higher with TAF compared to TFV.
Christian Callebaut and colleagues from Gilead created a large panel of reverse transcriptase mutation complexes derived from patients samples previously tested in the Monogram single-cycle phenotypic assay, including multiple combinations, with and without M184V, K65R, Q151M, T69ins and multiple TAMs and performed multi-cycle HIV infections to establish TAF activity against these viral isolates, Viral breakthrough experiments were subsequently performed, passaging HIV infected MT-2 cells every 4-5 days for 4 weeks to establish antiviral activity and related in vitro phenotypic cut-offs for TAF compared to TFV. The EC50 and EC95 values for each compound were used to establish intracellular IQ95 values, and target levels of 1 uM and 50 uM for TAF and TFV respectively.
TAF activity was highly inversely correlated with phenotypic fold change, loosing sensitivity when reaching 15-20 fold changes, with this requiring greater than three TAMs. The group also looked at two higher concentrations for each complex allowing an opportunity to see whether higher dosing might overcome further resistance.
Table 1 includes six of these complexes with the corresponding fold change in the single-cycle infection Monogram assay and mutli-cycle MT-2 cell exposure, and the time to viral breakthrough over 28 days if this occurred.
These in vitro results suggest that TAF is likely to be active against HIV that has developed resistance to TDF and other NRTIs and provide the first evidence for clinical advantages from the advanced formulation that are more than theoretical.
|Virus ID||Category||Mutation complex||FC Monogram||FC MT-2 cells||Time to breakthrough (days)|
|6||3 TAMS||M41L, L210W, T215Y||1.8||3.8||TAF >28 days||TFV 13 days|
|11||K65R||K65R, M184V||1.8||6.1||TAF >28 days||TFV 4 days|
|15||K65R/Q15M||M41L, A62V, K65R, T69ins, K70T, L74V, V75I, Y115F, F116Y, Q151M, M184V||2.4||3.3||TAF >28 days||TFV 8 days|
|30||T69ins||D67E, T69SSG||4.5||10.1||TAF >28 days||TFV 4 days|
|31||5 TAMs||M41L, D67N, T69D, L210W, T215Y, K219R||4.5||21.9||TAF 8 days||TFV 4 days|
|34||5 TAMs||M41L, D67N, L210W, T215Y, K219R||5.8||14.7||TAF 8 days||TFV 4 days|
These data support the importance of developing TAF as an individual drug to ensure a solid dataset with a wide range of ARVs. This should be in addition to the current development programme for TAF that so far prioritises coformulations and fixed dose combinations over the single compound.
Hopefully these results will encourage Gilead to recognise that future access to TAF should not be restricted to co-bundled formulation with other compounds owned by this company and its partners.
There will be an ethical urgency for patients with existing drug resistance in both wealthy and resource limited countries to be able to have access to this new formulation.
Callebaut C et al. Antiviral activity of tenofovir alafenamide (TAF) against major NRTI-resistant viruses: improvement over TDF/TFV is driven by higher TFV-DP loading in target cells. International Workshop on HIV and Hepatitis Virus Drug Resistance and Curative Strategies, 4-8 June 2013, Toronto. Oral abstract 23.
http://www.intmedpress.com/journals/avt/abstract.cfm?id=2658&pid=88 (abstract book)
http://www.informedhorizons.com/resistance2013/pdf/Presentations/Callebaut.pdf (PDF slides)