Systematic screening of viral and human genetic variation identifies antiretroviral resistance and immune escape link

Acceptance summary:

This paper uses a rigorous methodological approach to identify an interesting and novel mechanism of HIV drug resistance in addition to acquired and transmitted drug resistance: accidental resistance arising coincidentally due to immune escape. This finding is of potential clinical importance as such mutations can arise spontaneously during untreated HIV infection in the context of a specific HLA-type.

Decision letter after peer review:

Thank you for submitting your article "Systematic screening of viral and human genetic variation identifies antiretroviral resistance and immune escape link" for consideration by eLife. Your article has been reviewed by 2 peer reviewers, including Joshua T Schiffer as the Reviewing Editor and Reviewr #1, and the evaluation has been overseen by Jos van der Meer as the Senior Editor.

The reviewers have discussed their reviews with one another, and the Reviewing Editor has drafted this to help you prepare a revised submission.

Essential revisions:

1. Please expand and clarify the description of the DRM and genotyping data. Please specify the number of different HLA-types, the relative frequencies of the predominant HLA-types, the number of different drug resistance variants found in ART-naïve patients and their relative frequency in the population as well as frequencies of multiple DRMs.

2. In the results, please include a first paragraph that puts the subsequent analysis into context and describe the purpose of the cross-sectional and survival analyses before presenting the results.

3. Either eliminate the term "literature search" or specifically describe the search methodology that was used to perform this search.

4. Include viral load data if available as described by Reviewer 1.

Reviewer #1:

This paper by Nguyen and colleagues identifies 3 possible HIV mutations which are linked by HLA-association and drug resistance. The conclusions are justified based on the analysis. The results are unlikely to influence clinical practice as the identified mutations are of only occasional importance in the clinic. However, the general concept is of great relevance to HIV and other viral infections.

Strengths of the study are:

1. Novel conclusions: the paper is important because it identifies a new source of possible drug resistance in addition to selective pressure during failed therapy and transmitted drug resistance.

2. Methodology strength: the authors do a nice job of attempting to move towards causality rather than correlation. Specifically, they establish strength of correlation, temporality (the survival analysis showing accrual of the new mutation over time), assessment of possible confounding variables such as ruling out transmitted mutations with the survival analysis and looking at an interaction term with infection time, assessing mechanistic plausibility with literature review and utilizing a very large cohort.

3. Thoughtful and clearly written intro and discussion.

There were no major weaknesses with the study from my perspective.

This study was extremely clearly written and I believe the scientific conclusions are justified by the analysis. One area of interest would be viral loads among study participants with specific HLA/drug resistance pairs. If possible and if data is available, then it would be interesting to see whether onset of a new mutation as seen in the survival analysis is associated with a decrease in viral load due to a fitness cost. There would be a natural comparison to make versus viral load changes in all other participants, as well as those with relevant HLA types who do not develop new drug resistance mutations. A similar approach with CD4 T cell count trajectories would also be interesting and in theory easy to perform.

Reviewer #2:

Drug resistance mutations (DRMs) are known to emerge and be selected for in patients on ART experiencing treatment failure. Transmission of DRMs can lead to high levels of drug resistance in treatment-naïve individuals in resource-limited settings, which may compromise first-line treatment regimens. Here the authors investigated whether the emergence and persistence of DRMs in untreated individuals could also occur as a side effect of immune selection. To this end, they screened for associations between drug resistance mutations and different HLA-1 alleles in a large cohort of nearly 4,000 treatment-naïve patients.

The authors identified three DRM-HLA-1 pairs where the presence of the HLA-1 allele was predictive of the patient having acquired the DRM prior to starting ART. Similar analyses of other patients cohorts have also identified potential associations between DRMs and specific HLA-1 alleles, so in that sense, the novelty of these findings is limited. However, this study substantially extends previous work by incorporating additional analyses that account for the duration of infection before treatment and the time to emergence of drug resistance, leveraging their exceptionally detailed dataset. These analyses revealed that patients without the HLA-B18 allele who had acquired the DRM at transmission were likely to revert to wildtype over time due to the fitness cost associated with the mutation, whereas patients with the HLA-B18 allele either retained the transmitted DRM or generated it de novo to escape immune pressure. Longitudinal survival analysis further indicated that patients with the HLA-B18 and HLA-B35 alleles who did not initially have the associated DRMs were significantly more likely to acquire them than patients without the alleles.

Overall, the conclusions of this paper are well supported by data. The statistical methods used are rigorous and straightforward to apply to cohort data for other populations. However, the description of the DRM and genotyping data is minimal, and its presentation is somewhat confusing. While the three identified HLA-DRM pairs were analyzed extensively, it is not clear how frequent these genotypes and DRMs (independently of each other) are in the patient cohort. As currently written, the manuscript lacks context for assessing the contribution of immune-driven DRMs relative to transmission-acquired DRMs in the treatment-naïve HIV+ population. Overall, however, this paper provides a valuable contribution to the field as a proof-of-concept demonstration of the emergence of immune-derived DRMs.

The Results section would greatly benefit from a first paragraph that puts the subsequent analysis into context. Currently it is challenging to get any sense for the distribution of DMRs and HLA-types in the population. It would help to specify the number of different HLA-types, the relative frequencies of the predominant HLA-types, the number of different drug resistance variants found in ART-naïve patients and their relative frequency in the population. What proportion of the patients that were included in the study had any drug resistance mutations at all? Did any patients have multiple DRMs?

In my opinion, the Results section would flow more logically if the purpose of the cross-sectional and survival analyses were discussed before presenting the results. E.g. A cross-sectional analysis was performed to assess "…" and a survival analysis to assess "…".

Specifying "literature search" as an analysis methodology seems a little clunky. Why not simply state that the B18 HLA type has been shown to bind to the epitope containing the E138 DR mutation in several previous studies [listing citations]?

Would the cross-section analysis have more power if the two potential HLA alleles associated with the PR-E138 DRM were compared simultaneously against a reference group containing all other alleles? In the current analysis, the two potentially important alleles are essentially compared against each other (as part of the "other" category).

It would be helpful if the frequencies in Figure 3A would be spelled out in text rather than just shown in the table. E.g. what proportion of patients had the DRM-associated HLA alleles, and what fraction of these patients subsequently acquired the DRMs?